Odontogenic cyst is a common lesion that can happen after inflammation of the dental pulp. The therapeutic approach of these cysts is made at dentist's offices, and depending on their extension, they may develop oroantral fistula and chronic sinusitis. The objective of this study is to propose the videoendoscopic treatment of the odontogenic cyst with expression in the maxillary sinus. We made a retrospective study of four cases of cysts of dental origin, with intra-sinusal extension, complicated with oroantral fistula and chronic sinusitis of maxillary sinus after curettage in a dentist's office. We used the videoendoscopic technique through transmaxillary approach to access the intra-sinusal cyst. All the four patients presented resolution of the infectious manifestation and healing of the oroantral fistula, without recurrence within two years of follow-up. Videoendoscopic surgery is a safe and effective method for the management of odontogenic cysts with extension to maxillary sinus, and it may prevent oroantral fistula formation and chronic sinusitis. Odontogenic cysts are lesions that frequently rise in the maxilla and/or mandible, originating from epithelial remains associated with odontogenesis. The most frequent types are: periapical cyst (65%), dentigerous cyst (24%) and primordial or keratocyst (5 a 8%).2.The majority of these cysts are managed at dentist's offices using surgical approaches such as enucleation with use of Milton's solution or combined with cryotherapy, simple enucleation, curettage, marsupialization and tooth extraction.Intervention of cysts found in the maxillary sinus may lead to oroantral fistula formation and chronic rhinosinusitis. This study aims at proposing a videoendoscopic technique to treat odontogenic cyst with intrasinusal extension as a complementary approach of the odontological treatment.Four cases of dental cysts with intrasinusal extension were operated on  and 2. TPatients were surgically treated under general anesthesia. The procedure of choice was combined access through the canine fossa with lower meatotomy. Such approach is performed by means of a 10mm-circumferential antrotomy to allow simultaneous management of the optical fiber  and of small clamps , achieving cyst resection. The cyst should be removed in a way that only the bone bed remains on the sinusal wall at the implantation spot. In case of an oroantral fistula, curettage of the borders is also done. Occlusion is performed with a septal or conchal bone fragment as well as a flap rotation of the jugal or palatine mucosa  sutured Infections of all four patients were resolved and fistula healing was reached. No relapses were reported after 2 years of follow-up. Cyst fragments obtained during surgery were sent for clinical pathology analysis and the results were compatible to periapical cyst. No signs of malignancy were found in the analyzed cases.a)Epithelial fragments of the teeth crown.b)Epithelial debris of Malassez c)Epithelial debris of Seres (remains of teeth lamina)d)Fragments of teeth germ cells, including enamel, dental papilla and dental sac.A cyst is defined as a cavity layered by epithelium hosting liquid or semi-solid material originating from embryonic epithelial tissue. The epithelium of the odontogenic cyst may come from the following structures:The most accepted hypothesis for cyst development is proliferation of epithelial fragments with formation of islets. Due to its nonvascular aspect, degeneration occurs centrally. In addition, as it is distant from the adjacent connective tissue, release of enzymes leads to degeneration of its own cell protoplasm, and dead cells turn into liquid material. The intracystic liquid has higher osmotic pressure than that of surrounding tissues, which leads to progressive cyst growth.The dentigerous cyst occurs after development of the tooth crown with accumulation of liquid between the enamel epithelium and the tooth crown, directly linked to an unerupted tooth. This lesion frequently occurs in maxillary canine teeth and third lower molar teethRadiographically, this structure is characterized by a well-defined radiolucent image of bone cortical, involving an unerupted tooth crown and cervical onset. In order to differ from a dental sac image, the x-ray image should not be smaller than 2.5mm of extension. It may be central, lateral, circumferential or eruptionalCell degeneration of the enamel's starry reticulum gives rise to the primary cyst before the mineralized tissue is formed. This cyst appears in place of a normal, unerupted or supra-numerous toothThe periapical cyst, the most common type, is the result of an inflammatory stimulus promoted by the dental pulp. In general, they are asymptomatic, detected by x-ray, and occur in the third or fourth decade of life. They appear as a radiolucent image of homogenous, oval or round density, which is related to an intact radicular apex with rupture of the hard layer and a radio-opaque bone sclerosis liningWe observed that the videoendoscopic surgery is a feasible alternative to treat an odontogenic cyst with maxillary sinusal extension, which may be adopted simultaneously to prevent sinusal suppuration and complete cyst exeresis.Correction of oroantral fistulas caused by teeth extractions should be done by means of autologous bone graft and mucous flap rotation with suture.


The association between maternal obesity and preterm birth remains controversial and inconclusive, and the effects of gestational diabetes mellitus (GDM) and preeclampsia (PE) on the relationship between obesity and preterm birth have not been studied. We aimed to clarify the relationship between prepregnancy body mass index (BMI) and the phenotypes of preterm birth and evaluate the mediation effects of GDM and PE on the relationship between prepregnancy BMI and preterm birth.We conducted a prospective cohort study of 43,056 women with live singleton births from 2017 through 2019. According to the WHO International Classification, BMI was classified as underweight (BMI < 18.5 kg/m2), normal weight (BMI 18.5–25 kg/m2), overweight (BMI 25–30 kg/m2) and obese (BMI ≥30 kg/m2). Preterm birth was defined as gestational age less than 37 weeks . The clinical phenotypes of preterm birth included spontaneous preterm birth (spontaneous preterm labor and premature rupture of the membranes) and medically indicated preterm birth. We further analyzed preterm births with GDM or PE. Multivariable logistic regression analysis and causal mediation analysis were performed.Risks of extremely, very, and moderately preterm birth increased with BMI, and the highest risk was observed for obese women with extremely preterm birth . Maternal obesity was significantly associated with spontaneous preterm labor , premature rupture of the membranes  and medically indicated preterm birth . GDM and PE mediated 13.41 and 36.66% of the effect of obesity on preterm birth, respectively. GDM mediated 32.80% of the effect of obesity on spontaneous preterm labor and PE mediated 64.31% of the effect of obesity on medically indicated preterm birth.Maternal prepregnancy obesity was associated with all phenotypes of preterm birth, and the highest risks were extremely preterm birth and medically indicated preterm birth. GDM and PE partially mediated the association between obesity and preterm birth.The online version contains supplementary material available at 10.1186/s12884-022-04780-2. Preterm birth is a crucial global public health issue. Globally, preterm birth has been found to affect an estimated 10.6% of livebirths . PrematuMaternal obesity is a growing public health problem worldwide, and it is the most common medical condition in women of reproductive age. Obesity causes adverse consequences for both mothers and their children. Given the high prevalence and associated risks, maternal obesity has been identified as the most important preventable risk factor for adverse pregnancy outcomes in many countries.Prepregnancy obesity has been found to be associated with preterm birth; however, the relationship remains controversial and inconclusive in the literature. Some previous studies have found a positive correlation between obesity and preterm birth , 6, whilGiven the above controversial, inconclusive results and limitation, we performed a prospective cohort study to explore the relationships between prepregnancy BMI and the phenotypes of preterm birth. Maternal obesity has been found to be related to gestational diabetes mellitus (GDM) and preeclampsia (PE), and these two diseases are important maternal factors for preterm birth ; however+ 6 weeks who consented to participate in the cohort from 2017 to 2019. Postmature birth was excluded. All participants signed written, informed consent and agreed to be contacted for future studies. Our obstetric professional research assistants collected a comprehensive set of data on pregnancy including maternal characteristics, maternal risk factors and pregnancy complications at every antenatal clinic visit. After giving birth, delivery information and neonatal outcomes were abstracted from medical records. Quality control on this dataset included procedures for monthly quality checks and an annual secondary review of 10–15% of the records.Birth Cohort in Shenzhen (BiCoS), conducted by Shenzhen Maternity & Child Healthcare Hospital, is a prospective observational birth cohort study. We recruited participants with a singleton pregnancy at gestational age of 6–8A total of 43,056 pregnancies were available for inclusion in this study. We excluded 860 (2%) pregnancies from the dataset for the following reasons: missing maternal prepregnancy BMI, prepregnancy hypertension or diabetes, cervical incompetence, still birth and neonatal abnormalities.+ 6 gestational weeks) in light clothing and without shoes, and were obtained using a stadiometer and scale with calibrated electronic scale. Prepregnancy BMI was calculated using the above values. Information about GDM or PE was classified by the woman’s physician at the time of hospital discharge. We also included the following risk factors in the analysis: maternal age, education level, nullipara, parity and assisted reproductive technology.The primary explanatory variable was maternal prepregnancy BMI (weight in kilograms/height in meters2). We used the World Health Organization International Classification of BMI . The catPreterm birth was defined as delivery occurring before 37 weeks of gestation. Gestational age was based on the last menstrual period if the last menstrual period and the earliest ultrasound estimate were within 10 days of each other. If not, the earliest ultrasound evaluation was used to define gestational age. We subdivided the outcome into three groups by gestational age: extremely (< 28 weeks gestation), very (28–31 weeks gestation), and moderately (32–36 weeks gestation) premature . The cliP-value < 0.05.Continuous variables are reported as the mean ± standard deviation (SD) and were compared using Student’s independent t-test or Mann-Whitney U test depending on the normality assumption. Categorical variables are presented as numbers and percentages and were compared using the chi-square test or Fisher’s exact test . A multivariate logistic regression model was used to calculate odds ratios (ORs) with 95% confidence intervals (CIs) of BMI for preterm birth after adjusting for maternal age, educational level, and assisted reproductive technology. Relative risk (RR) was used to investigate the preterm birth risk in participants with or without GDM/PE. The above analyses were performed using IBM SPSS Version 25 . Furthermore, causal mediation analysis was used to estimate the OR for the natural direct effect (NDE) and the natural indirect effect (NIE) of preterm birth mediated through GDM or PE. We also estimated the proportion of the effect mediated to reflect the extent of mediation, where 100% indicates that all of the total effect is mediated and 0% indicates that there is no mediation. These analyses were performed by using R statistical software version 3.5.2 with the package ‘mediation’. The statistical significance level for all the tests was set at a two-tailed P < 0.001). Compared with women with full-term births, those with preterm births had a significantly higher rate of GDM or PE   births for the following reasons: missing prepregnancy BMI data (397), preexisting diabetes or hypertension (281), cervical incompetence (94), stillbirths and neonatal abnormalities (88). The final study population included 42,196 deliveries of live singleton infants, including 2768 (6.56%) preterm births. The average age of all participants was 31.26 ± 4.46 years, the prepregnancy BMI was 21.08 ± 2.84, and the parity was 0.60 ± 0.62. Most participants had a bachelor’s degree (65.49%), and only 1381 (3.27%) became pregnant through assisted reproductive technology. The participants who experienced preterm birth had a significantly higher maternal age and assisted reproductive technology rate and lower educational level . The same results were observed among women with PE  (Table SAs indicated in Table SP < 0.001) and PE ; preterm birth was positively associated with GDM  and PE .The direct association between BMI and GDM/PE, GDM/PE and preterm birth were also confirmed using logistic regression. It was found that maternal obesity was significantly associated with the incidence of GDM  and preterm birth. As indicated in Table Given the above results, causal mediation analysis was conducted to investigate the mediation effect of GDM or PE on the relationship between BMI  during which women have little change in their weight. Most importantly, we are the first to study the mediation effect of GDM or PE on the relationship between maternal prepregnancy BMI and preterm birth phenotype.Although some confounding factors were controlled, alcohol consumption and maternal smoking were not adjusted because only 10 women in our data reported a history of alcohol consumption and smoking. Therefore, we did not adjust for those two variables.However, there are a number of potential limitations of our study. First, we acknowledge that although we had a large cohort, the number of subjects was small in the obese category when we stratified preterm birth by gestational age and clinical phenotype. We were also unable to stratify the clinical phenotypes of preterm birth by gestational age. Similarly, when we conducted the mediation analysis, the sample size of each subgroup was limited. Additionally, although we attempted to adjust for potential confounding variables, we did not have information on variables such as other pre-existing medical conditions and previous history of preterm birth.In conclusion, our study notes that maternal prepregnancy obesity is an independent risk factor for all phenotypes of preterm birth. In addition, we conclude that GDM and PE partially mediate the association between prepregnancy obesity and preterm birth. Our findings support the potential importance of interventions to reduce prepregnancy obesity (such as reasonable diet and regular exercise) as an important strategy to reduce obesity-related pregnancy diseases and premature births.Additional file 1: Table S1. Maternal Pre-pregnancy BMI and Risks of Clinical Phenotypes of Preterm Birth without GDM/PE.Additional file 2: Table S2. The relative risks of preterm birth with or without GDM/PE.Additional file 3: Table S3. Distribution characteristics of GDM/PE in Different BMI Groups between Preterm Birth.


Data regarding mother and infant characteristics were collected. Gal-9 was present in a lower concentration range than Gal-1 and Gal-3 in plasma, specifically in UCP. A major finding in the current study is that Gal-1, -3 and -9 were detected for the first time in all the transitional breast milk samples and no differences were found when comparing the two breastfeeding time points. Finally, Gal levels were associated with some maternal and infant characteristics, such as gestational age, pregnancy weight gain, maternal diet, the gender, infant growth and infant infections. In conclusion, Gal levels seem to be involved in certain developmental aspects of early life.Galectins  are a family of conserved soluble proteins with high affinity for β-galactoside structures. They have been recognized as important proteins for successful pregnancy. However, little is known about their presence in breast milk and their role in early infancy. Gal-1, -3 and -9 concentrations were evaluated by Multiplex immunoassays in mother–infant pairs from the MAMI cohort in maternal plasma (MP) ( Galectins  were originally described as a group of proteins, produced by all cell types at varying levels, with high affinity to β-galactosides and high similarity in the sequence of the carbohydrate-binding sites, located in the carbohydrate-recognition domain (CRD) . Later oGals can have both immunosuppressive and immune regulatory functions, mainly depending on the cell type and the cell receptor they are binding to. Gal-1, -3 and -9 are some of the most abundant Gals, since they are expressed by a wide range of cells, including macrophages and epithelial cells . MoreoveLGALS1) was one of the most highly expressed in the decidua at term gestation [LGALS1 and LGALS3  had a higher expression in Cesarean section at term as compared to cases with spontaneous term labor, indicating an anti-inflammatory effect of Gals during pregnancy [The gene encoding for Gal-1  1 and Th17 cells. For this reason, a decreased expression of Gal-1 was positively correlated with failed pregnancies ,11,12. DLikewise, Gal-3 downregulation in the placenta, leading to placental dysfunction, was correlated with fetal growth restriction (FGR) . In addiGal-9 has been mainly related to the regulation of natural killer (NK) cells function to promote maternal–fetal tolerance in early pregnancy. Gal-9 levels are elevated during pregnancy as compared to healthy non-pregnant controls and decrease at parturition, having an immunoregulatory effect .Since breast milk (BM) contains abundant human milk oligosaccharides (HMO), which are a combination of the five monosaccharides glucose, galactose, fructose, N-acetyl-glucosamine and sialic acid , it seemDespite the increasing literature on the role of Gals on pregnancy, to our knowledge nothing is known about Gal-1, -3 and -9 presence in BM. For that reason, we evaluated the presence and concentrations of these Gals in human plasma and BM samples from two independent subgroups of mother–infant pairs within the MAMI cohort . The curz score was calculated with the WHO Anthro software (www.who.int/childgrowth/software/en/); accessed on 1 June 2020.A total of 15 healthy mother–infant pairs for plasma analysis, and an extended group of 23 healthy mother–infant pairs for BM analysis, were included into this sub-study from the MAMI cohort  . MaternaThe 140-item Food Frequency Questionnaire (FFQ) was used by a nutritionist to collect the maternal dietary records during the first week after birth. FFQ information was analyzed for the energy and daily intake of macro- and micronutrients, as commented in previous articles ,24. Regag for 10 min at 4 °C. The plasma was centrifuged again at 2500× g for 10 min at room temperature to deplete platelets in the plasma. The resulting supernatant was aliquoted and stored at −80 °C until further analysis, corresponding on the one hand to the maternal plasma (MP) and on the other hand to the umbilical cord plasma (UCP) [Maternal blood was obtained from 15 mothers and collected in sterile containers into anticoagulant (EDTA)-treated tubes immediately prior to delivery (20–30 min before expulsion or before incision in C-section deliveries), as in previous approaches . Arteriama (UCP) .g for 15 min at 4 °C. Finally, samples were frozen at −80 °C for latter analysis.BM samples for this study were obtained at 7 and 15 days after birth in the morning before lactation ,25. In bTM Multiplex immunoassay  using a Gal-1 Human ProcartaPlexTM Simplex Kit, a Gal-3 Human ProcartaPlexTM Simplex Kit and a Gal-9 Human ProcartaPlexTM Simplex Kit. To develop the analysis, the manufacturer instructions were followed as in previous studies [® analyzer, Luminex Corporation) at the Flow Cytometry Unit of the Scientific and Technological Centres of the University of Barcelona (CCiT-UB). The test interval was as follows: 394.46–615,700.00 pg/mL for Gal-1; 439.45–1,800,000.00 pg/mL for Gal-3 and 2.22–9100 pg/mL for Gal-9. One outlier UCP sample was discarded from the Gal study.The quantification of Gals  was performed by ProcartaPlex studies . The plahttp://www.rstudio.com/ (accessed on 20 April 2022)). Results are expressed as mean ± SEM, unless otherwise specified. Shapiro–Wilk and Levene’s tests were used to determine normality and homogeneity of data variance, respectively. When data were normally distributed, Student’s t test was used to assess significant differences between groups. When variables were not normally distributed, non-parametric tests were used, as Kruskal–Wallis and Mann–Whitney U tests. The spearman correlation coefficient was used to search for correlation between variables. Clustering of the individuals was analyzed by non-metric multi-dimensional scaling (NMDS) in Rstudio using the R package vegan (Community Ecology Package. R package version 2.4-6 (https://CRAN.R-project.org/package=vegan) (accessed on 11 October 2022) [p < 0.05 was considered significant for all analyses performed.Statistical analysis was performed using IBM SPSS Statistics for Windows, version 22  and RStudio version 4.2.0  (er 2022) . In the The assessment of Gal-1, -3 and -9 concentrations in maternal plasma (MP) and umbilical cord plasma (UCP) was performed in a subgroup of the MAMI birth cohort  consistiAll three Gals  were detected in both MP and UCP samples. Gal-1 and -3 were present in plasma at concentrations up to 60–80 ng/mL, while Gal-9 was present in a lower concentration range, up to 600 pg/mL. For Gal-1 and -3, no significant difference was found between the concentrations in MP and UCP A,B. ConvA Spearman correlation of pairwise comparison showed that Gal-1,-3,-9 concentrations were not correlated in mother–infant pairs, suggesting that the maternal–fetal transfer of Gals might not be the only factor involved in determining the UCP values .Maternal and infant factors  were assz score, antibiotic intake during pregnancy and/or exposure at the delivery day and neonate’s antibiotic intake and infections up to 12 months), gestational age and BMI z score at 6th month were significantly correlated with the NMDS ordination of the mothers based on Gal levels in MP   analysis  . NMDS anctively) .As for the effect of gender, a significantly higher concentration of Gal-9 in MP of mothers having male babies was observed .z score at 12 months and Gal-9 in UCP. A negative significant Spearman correlation was also found between infant antibiotic treatment during the first 6 months of life and Gal-9 concentrations in MP  and at day 15 (d15) from 23 mothers belonging to the MAMI cohort . The demInterestingly, Gal-1, Gal-3 and Gal-9 were all detected at both days, with Gal-9 being the Gal present at the lowest concentration . No sign2 = 0.3063, p value = 0.019) and the pregnancy weight gain tended to be correlated  , type of diet, gender of the baby and antibiotic intake during pregnancy and/or at the delivery day), none of them were significantly correlated with the NMDS ordination of the mothers based on Gal levels at day 7. However, the vector of the dietary component starch was highly related with the position of the individuals (envfit R= 0.065) . Despite= 0.065) . As for = 0.065) .Regarding the influence of perinatal variables on BM Gals at day 15, including dietary intake, no influence was found in the NMDS analysis . Howeverz score, antibiotic intake and infections), we also performed the NMDS analysis with the Gal levels at day 7 and day 15, separately. Multiple variables were identified as potential associated factors with the BM Gals at day 7   . In linectively) . Regardictively) , being sctively) C and Galctively) E.The role of galectins  in pregnancy and birth has gained interest in the past few decades, specifically, Gal-1, -3 and -9; however little is known about its relationship with some maternal, prenatal and postnatal factors. In the present study, we performed a characterization of these Gals in plasma at the delivery day, and in breast milk samples at two dates during the transitional breastfeeding period. To our knowledge, this is the first time that these Gals are found in breast milk and are associated with infant health parameters.In order to support a successful pregnancy, immune factors participate in coordinated communications between the mother and the fetus ,29. In tPlasma Gal levels have been reported to highly fluctuate in relationship to inflammatory conditions. He et al. (2016) reported similar levels for Gal-1, -3 and -9 after a large artery atherosclerotic stroke . EnningaThe finding that Gal-9 concentration in MP at birth was significantly higher for women giving birth to male fetuses than those giving birth to females corroborates previous findings . PreviouTo date, the presence of Gals in breast milk remains elusive, despite the physiological importance they could have in the infant health. On top of that, previous studies have reported an absence of Gals in human milk (from an unknown breastfeeding phase) when using western blot analyses  and whenRegardless of the similar Gal levels found at both time points, Gals at the beginning of the transitional stage, i.e., at day 7, showed stronger Spearman correlations to perinatal factors than day 15 samples. With regard to the maternal influencing factors, the overall dietary composition was not shaping the Gals profile however the highest influence due to diet was originated from starch and DPA intake. Early in life, a change in starch concentration in maternal diet has been related to a modulation of the lipid metabolism of newborn piglets  and a moz score during all the time period comprised from birth to 1 month. Previous studies have shown that multiple other milk components, such as fats, carbohydrates and proteins [With respect to the infant variables, the infant’s growth-related parameters were highly associated with the BM Gals concentrations, specifically to Gal-1 and 3 levels at day 7. The higher the Gal-1 and 3 at day 7, the lower the weight and BMI proteins ,49, leptproteins  and shorproteins  are relaThe joint analysis of all the factors influencing Gal-1, -3 and -9 concentrations in plasma and milk indicates that levels could be related with weight and immunity parameters. Interestingly, Gal-9 levels in MP at birth and in BM at day 7 postpartum are negatively correlated with infections and antibiotic intake during the first 12 months. While in BM a significant negative correlation was found for Gal-9 and infections during the first 12 months, in MP, the correlation was significant and negative for antibiotic intake during the first 6 months. These results suggest a possible role of Gal-9 in early life protection against infection.Comparison of Gal concentrations showed that Gal-1 concentration was about 1.5-fold lower in MP than in BM, Gal-3 was about 4-fold higher and Gal-9 was about 2.5-fold lower. A tolerogenic and anti-inflammatory effect has been associated with Gal-9 in pregnancy. The fact that it dropped significantly at 6 weeks postpartum could be in line with the fact that the concentration is lower in BM than in MP in our results, contributing to a higher Th1/Th2 balance needed to a correct immunological response in the infancy . All theThe limitations of this study include sample size and the absence of plasma-breast milk matching samples. These facts could have affected the statistical power and the strength of the associations with the perinatal outcomes, respectively, and deserve to be considered in the future.The results of the study demonstrated that Gal-1, -3 and -9 are present both in plasma at birth and in BM at transitional stage. Furthermore, Gal levels were not correlated in MP–UCP pairs, and Gal-9 was significantly lower in UCP with respect to the MP. Besides, Gal concentrations were similar when comparing both breastfeeding days corresponding to the beginning and the end of the transitional stage.Moreover, results showed that Gal levels were associated with some maternal and infant characteristics. Specifically, the gestational age and the gender were related to the MP Gal levels and the dietary starch and DPA on the BM Gal levels. Moreover, some other factors were also related, highlighting the infant antibiotic use with the MP Gal levels and the infant growth and infections with the BM Gals, specifically at the beginning of the transitional phase .Finally, the understanding of the presence of Gals in BM and the mother’s plasma during labor and the underlying molecular mechanisms could contribute to the design of interventions in order to sustain infant health and to prevent infant infections.


What are the associations between maternal milk feeding and cognitive, academic, and behavioral outcomes of preterm infants born at less than 33 weeks’ gestation?In this cohort study of 586 preterm infants, higher maternal milk intake during neonatal hospitalization was associated with higher performance IQ, better academic achievement in reading and math, and fewer attention-deficit/hyperactivity symptoms at 7 years of age. These associations persisted after adjustment for clinical and social confounders and were generally more pronounced among infants born at lower gestational ages.These findings affirm recommendations to provide maternal milk to hospitalized very preterm infants based on potential long-term benefits to neurodevelopment. This cohort study examines whether maternal milk feeding after very preterm birth is associated with cognitive, academic, and behavioral outcomes at school age and whether associations differ according to gestational age at birth, fetal growth status, and infant sex. Maternal milk feeding may have unique long-term neurodevelopmental benefits in very preterm infants.To examine the extent to which maternal milk feeding after very preterm birth is associated with cognitive, academic, and behavioral outcomes at school age.This prospective cohort study assessed 586 infants born at less than 33 weeks’ gestation at 5 Australian perinatal centers and enrolled in the Docosahexaenoic Acid for Improvement of Neurodevelopmental Outcomes study  who were evaluated at a corrected age of 7 years. The statistical analysis was completed on January 19, 2022.Maternal milk intake, including mean volume (milliliters per kilogram per day) during neonatal hospitalization and total duration (in months).Neurodevelopmental outcomes at 7 years of age were (1) IQ , (2) academic achievement , (3) symptoms of attention-deficit/hyperactivity disorder (ADHD) , (4) executive function , and (5) behavior .P values <.01).A total of 586 infants  born to 486 mothers  were included. Mean (SD) maternal milk intake in the neonatal intensive care unit was 99 (48) mL/kg daily, and mean (SD) maternal milk duration was 5.1 (5.4) months. Mean (SD) full-scale IQ was 98.5 (13.3) points. After covariate adjustment, higher maternal milk intake during the neonatal hospitalization was associated with higher performance IQ , reading scores , and math scores  and fewer ADHD symptoms . Longer duration of maternal milk intake was associated with higher reading , spelling , and math  scores. Maternal milk was not associated with improved full-scale IQ, verbal IQ, executive function, or behavior. Most associations were stronger among infants born at lower gestational ages, particularly less than 30 weeks (interaction In this cohort study of preterm infants, maternal milk feeding during the neonatal hospitalization and after discharge were associated with better school-age performance IQ and academic achievement and with a reduction in ADHD symptoms, particularly among infants born at less than 30 weeks’ gestation. Findings from studies of full-term infants are not directly generalizable to the very preterm population given substantial differences in postnatal nutrient requirements4 and neurodevelopmental risk profiles.5 Furthermore, the very preterm infant is fed expressed maternal milk during the several-months-long period from preterm birth to term equivalent age, a period of susceptibility to brain injury6 and one that coincides with an earlier stage of brain development than when the full-term infant is fed maternal milk. It is plausible that maternal milk may have different effects at different stages of brain development. Furthermore, within cohorts of preterm infants, the beneficial effect of maternal milk may vary by gestational age and fetal growth status as well as by infant sex.Compelling evidence links more prolonged and exclusive breastfeeding in infancy with improved neurodevelopmental outcomes in childhood.7 have linked maternal milk feeding during neonatal hospitalization with better neurodevelopmental outcomes in childhood. Differences in brain structure at term-equivalent age associated with maternal milk feeding in the interval from birth to hospital discharge provide further evidence to support a beneficial effect.11 Most studies7 of maternal milk and neurodevelopment have focused on outcomes at 2 years or earlier, in part because of the challenge and expense of retaining large cohorts for many years. However, many important neurodevelopmental domains are not reliably assessable until school age. To date, few studies have investigated the association between maternal milk feeding during neonatal hospitalization and neurodevelopment at school age, and existing studies7 are limited by small sample size, which precludes adjustment for multiple confounders and stratified analyses, as well as by a lack of detailed data on the dose of maternal milk to which the infant was exposed. In addition, previous studies6 examined a limited range of neurodevelopmental outcomes. Our aims were to examine the extent to which maternal milk feeding after very preterm birth is associated with cognitive, academic, and behavioral outcomes at school age and explore whether associations differ according to gestational age at birth, fetal growth status, and infant sex.Some but not all previous studies13 For this analysis, from the original cohort of 657, we excluded 20 participants who died before 7 years of age and 51 who were not evaluated at 7 years of age, leaving 586 participants. The sample size for each analysis ranged from 508  to 581  infants, depending on available outcome and covariate data. Institutional review boards of all 5 participating centers approved the original study, and parents provided written informed consent for their infant(s) to participate. This secondary data analysis was approved by the Women’s and Children’s Health Network Human Research Ethics Committee, the DINO Trial Steering Committee, and the Partners Human Research Committee. This study followed the Strengthening the Reporting of Observational Studies in Epidemiology (STROBE) reporting guideline.For this cohort study, we performed an observational secondary analysis of data from the Docosahexaenoic Acid for Improvement of Neurodevelopmental Outcome (DINO) randomized clinical trial. Briefly, 657 infants born at less than 33 weeks’ gestation without congenital anomalies were enrolled from January 1, 2001, to December 31, 2005, from 5 Australian perinatal centers. Details of recruitment, enrollment, and 7-year follow-up between 2008 and 2013 have been previously published.14 Pasteurized donor human milk was not used at any site during the study period.Once per week, study staff abstracted from the medical record the volume of maternal milk ingested by the infant and the infant’s body weight on that day. From those data, we calculated the mean volume of maternal milk normalized to body weight (milliliters per kilogram daily) fed to each infant during the entire hospitalization, including neonatal intensive care unit (NICU) and special care unit, as described previously.14At follow-up visits at term equivalent age and at 4, 12, and 18 months of corrected age, mothers were asked about the infant’s main source of milk  and the extent of maternal milk feeding . If the mother had stopped breastfeeding since the last visit, she was asked on which date she had stopped. To calculate the total duration of maternal milk feeding, including both direct breastfeeding and maternal milk fed via tube and/or bottle, we combined the duration of maternal milk intake while the infant was in the hospital with the duration reported at follow-up visits, as described previously.13 The primary outcome was general intellectual ability, as assessed by the full-scale IQ of the Wechsler Abbreviated Scale of Intelligence (WASI), administered by trained research personnel.15 For this secondary analysis, in addition to the full-scale IQ and verbal and performance IQ subscales, we were also interested in outcomes related to academic achievement and behavior, because these are domains in which preterm children perform more poorly than full-term children.18 To measure academic achievement in reading, spelling, and math, trained research personnel administered the Wide Range Achievement Test, Fourth Edition (WRAT-4).19 Parents reported symptoms of attention-deficit/hyperactivity disorder (ADHD) with the Conners Third Edition ADHD Index,20 behavioral problems with the Strengths and Difficulties Questionnaire (SDQ),22 and executive function with the Behavioral Rating Inventory of Executive Function (BRIEF).23 Both WASI and WRAT-4 are age standardized to a mean (SD) score of 100 (15). The Connors Third Edition ADHD Index T score is age standardized to a mean (SD) score of 50 (10); a normal score is less than 60, the elevated range  is 60 to 69, and a very elevated score  is greater than or equal to 70. BRIEF is age standardized to a mean (SD) score of 50 (10); a scale score or index greater than 64 is clinically indicative of dysfunction. In the SDQ, a total difficulties score greater than 13 indicates dysfunction. Higher scores on WASI and WRAT-4 indicate better performance, whereas lower scores on the reported scales from the Conners Third Edition ADHD Index, SDQ total difficulties, and BRIEF indicate better outcomes (fewer problems).Neurodevelopmental outcomes at 7 years’ corrected age for the DINO trial have been previously reported.Children who were too impaired to be tested were reviewed by a psychologist and assigned the lowest possible scores. Neither professional assessors nor parents were aware of the hypothesis for this secondary analysis. Assessors were unaware of infant feeding history and were also unaware of the child’s gestational age at birth, although they were aware that the trial enrolled children born at less than 33 weeks’ gestation and in theory could have calculated an individual child’s gestational age at birth from the child’s date of birth and estimated due date or corrected age. Parents could not feasibly be masked to infant feeding or gestational age at birth.z scores and percentiles using the Fenton reference24 and classified infants as small for gestational age (SGA) if the birth weight was less than the 10th percentile.On enrollment, mothers were interviewed about social factors, including income, educational level, occupation, race, smoking status, alcohol use, and the number of people living in the home. Study staff extracted data from the medical record regarding maternal parity and antenatal steroid exposure and infant sex, birth weight, gestational age, multiple birth status, and diagnoses, including necrotizing enterocolitis, bronchopulmonary dysplasia , sepsis, and severe intraventricular hemorrhage (grade 3 or 4) or periventricular leukomalacia. We derived birth weight for gestational age 26 Confounders adjusted for in the models included maternal educational level, maternal occupation, number of adults and children living in the home, maternal smoking, maternal alcohol use, parity, race, birth weight (continuous), gestational age (continuous), plurality, exposure to antenatal steroids, treatment group (high vs standard docosahexaenoic acid), and center. We conceptualized maternal race as a social construct. We did not adjust for infant diagnoses because they occurred after the start of measurement of the exposure variables and therefore may be on the causal pathway rather than confounders. We assessed effect modification of the exposure variables (linear continuous) by gestational age (linear continuous), fetal growth status (SGA vs not SGA), sex , and treatment group (high docosahexaenoic acid vs control). When looking at effect modification by SGA, we removed birth weight and gestational age as confounders because SGA is defined as birth weight less than the 10th percentile for gestational age. Regarding effect modification for gestational age, we do not report estimates for gestational age categories with fewer than 5 participants. A 2-sided P < .05 was considered to be statistically significant. We used R software, version 4.0.3  for all analyses.27 The statistical analysis was completed on January 19, 2022.Analyses followed a prespecified statistical analysis plan. The exposure variables were mean maternal milk intake volume and duration of maternal milk intake up to a corrected age of 18 months. The main outcome variables were WASI full-scale, verbal, and performance IQ scores; WRAT-4 reading standard, spelling, and math computation scores; ADHD symptoms (Conners Third Edition ADHD Index T score); BRIEF global executive composite score; and SDQ total difficulties score. To estimate the association of the human milk intake with neurodevelopmental outcomes at 7 years of age, we fitted linear models using generalized estimating equations with an independence working correlation structure to account for nonindependence of multiple births and repeated pregnancies and adjusting for clinical and social variables that we conceptualized as potential confounders.From the DINO trial’s original cohort of 657 infants, we excluded 20 participants who died before 7 years of age and 51 participants who were not evaluated at 7 years of age, leaving 586 participants. The 20 infants who died were born at a median gestational age of 25.4 weeks . Of survivors, gestational age at birth was similar between participants who were vs were not evaluated at 7 years . Mothers of participants who were evaluated were more likely to have a tertiary education (245 [51.0%] vs 12 [31.0%]) than mothers of participants who did not participate.Characteristics of the 586 infants and 486 mothers included in the present cohort are shown in P for interaction = .04). Regarding SGA status (P for interaction = .03). There was no evidence of effect modification by infant sex  when multiplied over the typical daily intake in the NICU, which is 160 to 180 mL/kg daily. In addition, we found that the beneficial associations of maternal milk feeding with neurodevelopment were stronger for infants born at the lowest gestational ages.7 and are therefore limited given the substantial evolution in cognitive performance that occurs in very preterm–born children over time,29 as well as the relatively small range of cognitive and behavioral domains that are testable in young children. A few previous studies have reported school-age outcomes, but most lack detail on the dose and/or timing of maternal milk feeding, an important limitation given the typical decrease in maternal milk provision after the first month in the NICU30 and given evidence that the NICU hospitalization is a particularly sensitive period for preterm brain development related to nutritional exposures.31Our findings substantially extend previous literature on the neurodevelopmental benefits of maternal milk feeding for preterm infants. Of relevance to current practice are studies conducted during the era of routine human milk fortification. Most of those studies report outcomes in infants and toddlers32 A US study33 of more than 400 infants with very low birth weight (<1500 g) found that parent-reported feeding (direct breastfeeding vs no direct or expressed milk feeding) was associated with better visual motor integration at age 6 to 8 years, but associations with other measures were attenuated with full adjustment for confounders. A previous study8 in a different Australian cohort quantified the number of days on which maternal milk comprised greater than 50% of enteral intake during the first month of life and found positive associations with IQ, mathematics, working memory, and motor function tests at 7 years of age. We also found that a longer duration of maternal milk feeding, including the period after hospital discharge, was associated with benefits for academic achievement in terms of reading, spelling, and math scores, although the effect sizes were more modest than for the intake of maternal milk during the hospitalization. These findings are consistent with previously reported literature.32 Taken together, the available evidence supports the concept that a higher dose of maternal milk during the NICU hospitalization, and possibly after discharge, is associated with beneficial effects at school age on intelligence and academic achievement, particularly in mathematics given the consistency in this association across 2 separate cohorts.Consistent with our study, in a large, regional French cohort born in 1997, maternal milk feeding, compared with formula feeding, assessed at the time of NICU discharge, was associated with a protective effect against low IQ at age 5 years.34 Data linking maternal milk or breastfeeding with symptoms of ADHD in the preterm population are sparse, but our findings are consistent with a German Neonatal Network study35 in which parent-reported breastfeeding for 3 months or longer was associated with lower conduct and inattention/hyperactivity scores, indicating fewer problems in these domains, as assessed by the SDQ. Our finding of no association between duration of maternal milk intake and the SDQ inattention/hyperactivity scores may reflect the poorer sensitivity of the SDQ in detecting ADHD symptoms compared with the Conners Third Edition ADHD Index.36 Several studies in full-term populations have linked breastfeeding duration with ADHD diagnosis and/or related symptoms,39 whereas other studies, including a large randomized clinical trial of breastfeeding promotion,40 have reported no association.41 Notably, generalizability from studies of full-term infants to very preterm infants is limited given differences in timing of exposure with respect to underlying brain developmental processes42 and given the increased underlying risk of behavioral difficulties in preterm-born children.43 Overall, cumulative evidence to date suggests a possible protective effect of maternal milk against ADHD symptoms at school age in very preterm-born children, but firm conclusions are limited because of a lack of data from very preterm cohorts.In addition to associations with IQ and academic achievement, we noted that a higher dose of maternal milk was associated with lower Conners Third Edition ADHD Index T scores, indicating fewer symptoms of ADHD as reported by parents. Very preterm infants are at heightened risk of ADHD relative to their peers born at full term.45 Typically, attention problems also impact cognitive testing but often less markedly than school performance.46The findings regarding ADHD symptoms may also help to explain another observation—that associations of maternal milk were stronger and more consistent with academic achievement than with IQ. It is plausible that an underlying attentional vulnerability may manifest as impaired school performance, independent of IQ, as has been observed in both general and very preterm cohorts.47 Maternal milk feeding during NICU hospitalization has been linked in several studies11 with differences in brain structure on magnetic resonance imaging at term-equivalent age, suggesting short-term effects during a critical period in development. Insights from animal models and human studies that link variation in exposure to specific milk nutrients and/or nonnutrient bioactive factors, or mixtures of these nutrients and bioactive factors, with early brain development and neurodevelopmental outcomes can inform randomized clinical trials and eventually new clinical strategies for supplementation.48A beneficial association of maternal milk with school-age neurodevelopmental outcomes is biologically plausible when framed within the Developmental Origins of Health and Disease paradigm and in the context of mounting research describing the myriad, potentially beneficial nutrient and nonnutrient bioactive factors in human milk.This study has several strengths, including the large sample size, which allowed us to adjust simultaneously for multiple covariates and to perform analyses stratified by gestational age. High retention of the cohort through 7 years of age further strengthens internal validity. The study also has limitations. The main findings were robust to adjustment for potential medical and social confounders, but as with all observational studies, this study is limited by the potential for residual confounding by unmeasured factors in the NICU or home environment. The use of developmental therapies may have differed by feeding status, but these differences would be on the causal pathway and therefore should not be adjusted for. Given the large number of outcomes, some statistically significant findings may be spurious, although all were in the hypothesized direction, and 95% CIs provide information about the precision of estimates. Generalizability to very preterm infant populations outside Australia, particularly infants born in low-resource settings, may be limited, and racial and ethnic diversity of the cohort is also lacking.46 In addition, these findings suggest that hospital-based and societal efforts to increase maternal milk feeding through policies that protect, promote, and support lactation for women who deliver very preterm are well justified based on long-term benefits to child neurodevelopment.In this cohort study, we found that a higher dose of maternal milk during the critical period from very preterm birth to term-equivalent age was associated with improved cognitive and academic outcomes and fewer ADHD symptoms at school age. These associations were stronger for infants born before 30 weeks, suggesting that infants who are less mature at birth may derive greater benefit from maternal milk feeding after birth. These findings affirm recommendations by national and international organizations that fortified maternal milk should be the primary diet for infants born very preterm.


Here we use program data to create an empirical model predicting the year of attaining global elimination of TF1–9.Great progress has been made toward the elimination of trachoma as a public-health problem. Mathematical and statistical models have been used to forecast when the program will attain the goal of the elimination of active trachoma, defined as prevalence of trachomatous inflammation—follicular in 1–9 year olds  and baseline prevalence category. Ethiopia IUs had significantly different rates of reaching the TF1–9 elimination threshold after a trachoma impact survey (TIS) compared to non-Ethiopia IUs across all baseline categories. We used those estimates to predict when remaining active trachoma-endemic IUs (TF1–9 ≥5%) would have their last round of mass drug administration (MDA) based on the mean number of years required and number of MDA rounds already completed. Our model predicts that elimination of TF1–9 will be achieved in 2028 in Ethiopia (95% CI: 2026–2033) and 2029 outside of Ethiopia (95% CI: 2023–2034), with some IUs in East Africa predicted to be the last requiring MDA globally.We calculated the mean number of years (95% CI) observed for an implementation unit (IU) to move from a baseline TF1–9 elimination is realistic with the caveat that although disease elimination progress can be predicted for most IUs, there is an important minority of IUs that is not declining or has not yet started trachoma elimination activities. These IUs represent an important barrier to the timely global elimination of active trachoma.Our empirical estimate is similar to those resulting from previous susceptible-infectious-susceptible (SIS) and mathematical models, suggesting that the forecast achievement of TF Mathematical and statistical models have been used to forecast the global decline of active trachoma prevalence as a result of interventions against the disease. Here we used trachoma program data to create an empirical model predicting the year of attaining global elimination. We analyzed over 20 years of trachoma implementation and survey data to calculate the mean number of rounds of mass drug administration required for implementation units to fall below the elimination threshold for active trachoma. We then compared these means against the number of rounds of mass drug administration previously undertaken for all areas still above the elimination threshold to produce a forecast of the last year of antibiotic distribution for each implementation unit. Our model predicts that elimination will be achieved in 2028 in Ethiopia (95% CI: 2026–2033) and 2029 outside of Ethiopia (95% CI: 2023–2034), with areas in East Africa predicted to be the last requiring mass drug administration globally. However, although disease-elimination progress can be predicted for most implementation units, there is an important minority of implementation units that is not declining or has not yet begun implementation. These areas represent an important barrier to the timely global elimination of active trachoma. Achieving and sustaining both goals, along with the existence of a system able to identify and manage incident TT cases, evidences the elimination of trachoma as a public-health problem [1–9 ≥5% [1–9 ≥ 5%) has decreased year-on-year since a global peak in 2015 [In 1996, the World Health Organization (WHO) and partners launched the WHO Alliance for the Global Elimination of Trachoma by 2020 (GET2020), which aims to reduce the prevalence of: 1) trachomatous inflammation—follicular in 1–9 year olds , with a TF1–9 between 5–9.9% receiving one round, a TF1–9 between 10–29.9% receiving three rounds, a TF1–9 between 30–49.9% receiving five rounds, and a TF1–9 ≥50% receiving up to seven rounds of MDA [1–9 <5%) and therefore inform if MDA can be stopped.Once-yearly mass drug administration (MDA) of oral azithromycin is a key component of the WHO-recommended surgery, antibiotic, facial cleanliness, and environmental improvement (SAFE) strategy for trachoma elimination as a public-health problem (EPHP). The number of years of MDA recommended is dependent on the TFs of MDA . After rChlamydia trachomatis), there is a risk of recrudescence. In order to monitor for recrudescence, trachoma surveillance surveys (TSS) are conducted in formerly endemic IUs at least 2 years after the TF1–9 prevalence is demonstrated to be <5% and where there have been no additional rounds of MDA [1–9 <5% at TSS, a prevalence of TT in adults unknown to the health system <0.2%, and satisfied all other requirements, a country may submit a dossier to the WHO to be validated as having attained EPHP. By 2021, eleven countries had been validated by WHO as having achieved trachoma EPHP [Since elimination as a public-health problem is not the same as elimination of transmission of the disease-causing organism  in partnership with theWHO. The database contains data on trachoma prevalence surveys and reported SAFE implementation activities. While there is some variation in the sampling design and survey methodology for any given survey result, the majority of surveys since 2013 followed a standardized methodology to measure TF Project  and its  Project . An IU i Project . Because Project . Countri1–9 without presence of infection: Fiji, Papua New Guinea, Solomon Islands, and Vanuatu [1–9 <5%) were also excluded. Countries were grouped not by WHO region, but by geographical groupings sharing similar epidemiology.Survey data determined to be historic (a baseline followed by another survey in the absence of implementation) were excluded. Data from the following Pacific Islands were excluded due to atypical TF Vanuatu ,19. IUs 1–9 category, and the proportion of IUs with a TIS reaching the elimination threshold. The null hypothesis was that the proportion of IUs with a TIS above the elimination threshold was the same in Ethiopia vs. non-Ethiopia for all baseline categories.For IUs with a TIS, a Cochran–Mantel–Haenszel test was performed to assess the relationship between region (Ethiopia vs. non-Ethiopia), baseline TF1–9 <5%), we took the mean of the total number of rounds of MDA required for all IUs that have reached the elimination threshold as of July 2021 based on the Ethiopia/non-Ethiopia status and baseline TF1–9 prevalence category. All IUs currently under the elimination threshold were included in this calculation, regardless of survey history . Then, for IUs with TF1–9 ≥5% as of July 2021, we subtracted the number of rounds already conducted in an IU since the most recent survey from the mean number of rounds projected to be required to reach the TF1–9 threshold in order to calculate the number of rounds remaining in each IU.To calculate the estimated number of rounds of MDA required to reach the elimination threshold (TFWe used the number of rounds remaining (rounded up to the nearest whole number) to project the last year of MDA required per IU, assuming one round of MDA per year. The 95% confidence intervals were calculated as ±1.96 times the standard deviation . The minimum last year of MDA was set to 2021 to account for cases where either the required number of rounds has already been reached, or where the lower bound of the 95% CI was negative and the projected last year of MDA was prior to 2021.For cases in which there was ambiguity in the data as to whether a survey or MDA occurred first , the month of MDA (where available) and intervention status at time of Zithromax application were used to place MDAs within a cycle. In cases where this information was not available, we assumed the MDA occurred prior to the survey, because this is more conservative for forecasting.For the analyses using geographic groups, countries were assigned to a category according to 1–9 ≥5%), of which the majority (60.4%) were in Ethiopia.A total of 2,082 IUs contributed data for 4,457 surveys, of which 2,364 were TIS. Nearly half of IUs (44%) were above the elimination threshold  as the number of currently known endemic IUs remaining by year Figs  and 2; bic group , and c) ic group .1–9 <5% and will no longer require MDA. The forecast mean last year of MDA for the global program is 2029 (95% CI: 2026–2034). Using the mean forecast, we anticipate that more than 50% of IUs currently above the elimination threshold will achieve elimination by 2024, with over 90% of the currently endemic IUs moving below the TF1–9 elimination threshold by 2026 . Programs should keep this in mind when allocating resources and planning realistic implementation timelines.1–9 <5% has been sustained in the absence of the mass distribution of antibiotics. Data suggest that around 9% of IUs show an increase in TF1–9 above the threshold at their TSS [Our forecast relies on several assumptions. We recognize that some of these assumptions, such as that there will be no interruptions in SAFE programming, are optimistic. Additionally, in accordance with current WHO guidelines, once IUs have reached the elimination threshold, MDA stops and the IU moves into the surveillance phase, with any necessary surgical interventions, facial cleanliness and environmental improvements continuing. After at least two years without antibiotic pressure, a TSS is conducted to determine whether TFheir TSS  and mustheir TSS . Therefo1–9 <5%. Operational research and clinical trials have demonstrated that more-frequent-than-annual MDA may accelerate progress to elimination [Additionally, this model assumes that there will be no technological or methodological advances in programming that accelerate achievement of the TFmination –24. A WHmination  in EthioThe COVID-19 pandemic affected the provision of health services worldwide, including MDA. Modelling work has indicated that, of the preventative chemotherapy NTDs, trachoma was one of the diseases most likely to "bounce back" the quickest, especially in high-transmission areas, and therefore catch-up MDA strategies in these settings are recommended, with such strategies having the potential to accelerate the timeline to elimination . In 20221–9 prevalence. Based on recent mapping results [Another assumption that we make is that there will be sufficient resources available  to implement SAFE and sustain elimination, which requires multi-sectoral partnerships and continued resource allocation before, during, and after achievement of elimination goals . However results , the vasSeveral of the Pacific Islands  are excluded from this analysis. Based on operational research  and foll1–9 <5% at an impact survey; therefore, it is difficult to predict the timeline to elimination for IUs that are behaving differently , since treatment is still underway and we do not yet know how many additional rounds of MDA will be needed. Some IUs have had 10 rounds of MDA or more and still have not yet reached TF1–9 <5% at TIS [1–9 <5% to stop MDA and sustain that progress for >2 years. Over time, these outliers will become increasingly important in determining global program progress. In these IUs, elimination remains more a question of "if" than "when"; thus research into risk factors for a TIS or TSS TF1–9 ≥5% and novel ways to address the disease in these areas will be a vital component of the global trachoma elimination strategy.An inherent limitation of all models is that they assume that the future will behave like the past. In order to be included in our calculation of the mean number of rounds of MDA required to reach the active trachoma elimination threshold, IUs have to have already reached TF% at TIS . Althoug1–9 elimination threshold between now and 2024 and an additional 40% estimated to reach this threshold by 2026. However, because global elimination requires sustained elimination in 100% of IUs, we have less confidence in our prediction of how much longer those trailing IUs will take to achieve TF1–9 elimination, which is why we need to start work now to make progress in those areas. In a global program, we must work everywhere, so by starting interventions where possible and exploring alternative treatment strategies that may accelerate progress in areas where intervention is not currently possible or where standard intervention does not appear to be working, we can achieve the goal of global elimination of trachoma.We can be reasonably certain that in the coming years (2022–2026) there will be a great deal of progress towards elimination of active trachoma, with over 50% of IUs forecast to achieve the TFThe views expressed in this article are the views of the authors alone and do not necessarily reflect the decisions, policies or views of the institutions with which they are affiliated, or of the funding agencies.


The brain glymphatic system is involved in the clearance of misfolding α-synuclein, the impaired glymphatic system may contribute to the progression of Parkinson’s disease (PD). We aimed to analyze the diffusion tensor image along the perivascular space (DTI-ALPS) and perivascular space (PVS) burden to reveal the relationship between the glymphatic system and PD. A cross-sectional study using a 7 T MRI of 76 PD patients and 48 controls was performed to evaluate the brain’s glymphatic system. The DTI-ALPS and PVS burden in basal ganglia were calculated. Correlation analyses were conducted between DTI-ALPS, PVS burden and clinical features. We detected lower DTI-ALPS in the PD subgroup relative to controls, and the differences were more pronounced in patients with Hoehn & Yahr stage greater than two. The decreased DTI-ALPS was only evident in the left hemisphere in patients in the early stage but involved both hemispheres in more advanced PD patients. Decreased DTI-ALPS were also correlated with longer disease duration, higher Unified Parkinson’s Disease Rating Scale motor score (UPDRS III) and UPDRS total scores, as well as higher levodopa equivalent daily dose. Moreover, the decreased DTI-ALPS correlated with increased PVS burden, and both indexes correlated with PD disease severity. This study demonstrated decreased DTI-ALPS in PD, which might initiate from the left hemisphere and progressively involve right hemisphere with the disease progression. Decreased DTI-ALPS index correlated with increased PVS burden, indicating that both metrics could provide supporting evidence of an impaired glymphatic system. MRI evaluation of the PVS burden and diffusion along PVS are potential imaging biomarkers for PD for disease progression. The pathological hallmarks of PD are a loss of dopaminergic neurons in substantia nigra pars compacta, and intracellular inclusions containing aggregates of α-synuclein2. The aggregation and abnormal deposits of insoluble α-synuclein in the central and peripheral nervous systems3 exacerbate the process of neurodegeneration4. Impaired α-synuclein clearance system has also been reported in PD, and could be considered as part of the PD pathology5.Parkinson’s disease (PD) is a common neurodegenerative disease that manifests with both motor and non-motor symptoms6. cerebrospinal fluid (CSF) enters from the para-arterial space into the brain parenchyma. Convective interstitial fluid (ISF) bulk flow is formed to propel waste products towards perivenous space. Subsequently, the waste product is drained out of the brain through the cervical lymphatic system8. Recent evidence emphasizes the role of the glymphatic system in eliminating neurotoxic protein aggregations in neurodegenerative diseases, such as amyloid-β in Alzheimer’s disease (AD) and α-synuclein in PD15. Glymphatic clearance dysfunction may lead to damage to the dopaminergic neurons secondary to increased α-synuclein deposition. Previous studies have provided more evidence that the blockage of the glymphatic pathway, which leads to drainage dysfunction, could aggravate α-synuclein pathology and exacerbate motor and memory deficits16. The glymphatic clearance dysfunction has been considered a possible contributing factor to abnormal α-synuclein aggregation in PD15. Conversely, aggregations of misfolding proteins may accumulate around blood vessels and block the glymphatic pathway18. Hence, PD could be considered a form of “central nervous system interstitial fluidopathy”, a concept used to describe diseases or conditions in which abnormal interstitial fluid dynamics are an important factor19. Therefore, evaluating the overall function of the glymphatic system could shed light on the pathophysiological mechanisms of PD20.The glymphatic system is a waste drainage system in the brain, involving the fluid interchange between the interstitial space and perivascular space (PVS)21, as well as with intrathecal contrast medium-enhanced MRI22 and dynamic PET23 in humans. In human studies, the invasiveness and other related risks limited the application of these methods. An improved and non-invasive method to reliably evaluate the function of the glymphatic system is essential.The glymphatic system has been studied using ex vivo fluorescent microscopic and in vivo two-photon imaging in animal models25. PVS burden in basal ganglia (BG) has been shown to correlate with the severity of PD motor symptoms27. PVS is a microscopic tubular structure that contains freely moving water molecules28. DTI-derived features might be influenced by structural alterations in the glymphatic system. Recently, a non-invasive method, DTI analysis along the perivascular space (DTI-ALPS), has been developed to evaluate the function of brain’s glymphatic system6. Water diffusivity along the x, y, and z axes of the periventricular white matter is generated based on diffusion sequences to calculate the DTI-ALPS index. The DTI-ALPS index represents the water diffusivity along the x-axis, which is parallel to the direction of PVS and could partially reflect the activity of the glymphatic system29. This method has been applied in a series of neurological diseases, including AD30, PD31, isolated rapid eye movement sleep behavior disorder32, epilepsy33, idiopathic normal pressure hydrocephalus34, etc. A lower DTI-ALPS index was correlated with increased severity of cognitive impairment in both AD30 and PD29. PD patients showed lower DTI-ALPS index when compared to normal controls29, as well as to people with other neurological conditions such as essential tremor31. These findings suggest the potential of the DTI-ALPS index as a non-invasive imaging biomarker in evaluating the integrity of the glymphatic system. Ultra-high field 7 T MRI, with increased spatial resolution, contrast and signal-to-noise ratio, improved the visualization and detection rate of PVS, especially small-sized PVS that might be neglected using conventional MRI technology35. Moreover, for DTI studies, 7 T MRI enables visualization of the white matter orientations and making sharp turns into the cortex, which are more difficult to see with the 3 T scanner36. The improved higher resolution and reduced partial volume effects make 7 T MRI superior in evaluating DTI-ALPS when compared with other investigations using clinical MRI scanners (1.5 T or 3 T).Recent advances in imaging techniques, including diffusion tensor imaging (DTI), make it possible to assess gross and microstructural changes in vivo. Such assessment in relation to clinical parameters can potentially serve as biomarkers to monitor disease progression. Since the PVS system is an important part of the glymphatic system, previous studies evaluated the glymphatic system indirectly by assessing PVS burdenThe current study aimed to investigate the integrity of the glymphatic system in different stages of PD using potential indicators, including the DTI-ALPS and PVS number/volume indexes derived from ultra-high field 7 T MRI. We evaluated these indexes to test the hypothesis that the diffusivity of perivascular fluid was reduced, and the perivascular structural impairment was increased in PD, which might represent glymphatic dysfunction and structural abnormality, respectively. We also hypothesized that both motor and non-motor symptoms might correlate with the progression of glymphatic impairment. Thus, we planned to further explore the correlation patterns of DTI-ALPS and PVS burden indexes with clinical profiles. Specifically, our study will focus on the role of these biomarkers that reflect the integrity of the glymphatic system, and explore whether impairment of the glymphatic system is correlated with the pathophysiological mechanism of PD.p = 0.03). The PDb group also had significantly higher HAM-D and HAM-A scores than those in the HCb group . Furthermore, there was no significant difference in the Fazekas scores between the PD and HC groups.Detailed demographic and clinical characteristics of the PD and healthy control (HC) participants are shown in Table p = 0.04), while no significant difference was found in the right hemisphere   Table . The difup Table .Based on receiver operating characteristics (ROC), the composite score combining the left/right DTI-ALPS and left/right PVS number/volume indexes yielded the best discriminatory performance in distinguishing PDb patients from their age- and sex-matched HCs with the area under ROC curve (AUC) of 0.85, while the AUC of the composite score for distinguishing PDa from HCs was 0.72 Fig. . When us37, HAM-A, and HAM-D scores  and volume (p = 0.001) in the right BG were significantly higher in PDb patients compared to HCb . Although only the PVS number in the left BG was significantly related to the total Fazekas score , PVS burden indexes showed a trend of positive correlation with the Fazekas score.To investigate whether the alteration of the DTI-ALPS index was consistent with increased PVS burden in PD, the correlations between the two metrics were calculated in all PD patients Table . PVS numHCb Fig. . SignifiThe DTI-ALPS index is a recently established non-invasive method to evaluate brain glymphatic function. In this study, we utilized higher resolution 7 T MRI, a superior tool in evaluating fiber tracks for DTI-ALPS and detecting small/normal-sized PVSs. We detected lower DTI-ALPS in both PD subgroups compared to age- and sex-matched HCs. We also observed a left/right differentiation of the DTI-ALPS index in PD patients, and a possible progression impacting bilateral hemispheres as the disease progresses. The lower DTI-ALPS indexes were correlated with increased disease severity, as reflected by UPDRS and LEDD. Furthermore, PVS burden was inversely correlated with DTI-ALPS indexes. Therefore, the DTI-ALPS indexes could be mutually complementary with the PVS burden, which could be used to evaluate the function of the glymphatic system.31. Although the DTI-ALPS was established to be measured in the periventricular space and evaluated the diffusion aberrations near PVS, it can represent the overall glymphatic function to some extent31.This study investigated the brain glymphatic system in PD with both DTI-ALPS and PVS burden indexes derived from 7 T MRI, and analyzed bilateral hemispheres separately. The DTI-ALPS index is calculated as the ratio of the diffusion in the perivascular direction along the medullary veins to the diffusion that is perpendicular to the principal fiber track direction29. In our study, only left-hemispheric glymphatic impairment was evident in the early disease course, while both hemispheres showed impairments in the PDb group as the disease advanced (H&Y >2). However, the left-hemispheric DTI-ALPS changes were more pronounced. This may suggest that glymphatic impairment starts from the left brain and propagates to the right through the progression of PD, although it potentially relates to the right hemibody being the location of motor symptom onset in nearly 70% of PD patients in this study. As the disease advances, such changes gradually progress to involve both hemispheres. This might be the reason that no significant differences were observed in DTI-ALPS indexes between right-onset and left-onset PD patients in our study cohort , lacunas, and microbleeds were found to be associated with DTI-ALPS index31, and its concomitant higher PVS burden suggests possible blockage of the glymphatic pathway. In PD, the failure of perivascular clearance may have led to α-synuclein deposition and the formation of a Lewy body49. Recent evidence has provided further insight into the role of the glymphatic system and its related pathways in neurodegenerative diseases. Since the waste clearance activity in the brain increases by 60% during the sleep state8, sleep disturbances might lead to suppressed glymphatic function. Thus, investigating the relationship between the brain’s glymphatic system and sleep could be meaningful in understanding the interstitial fluidopathy of PD. As the strongest prodromal marker of α-synucleinopathy disease, rapid eye movement sleep behavior disorder exhibited decreased DTI-APLS, indicating the presence of glymphatic system dysfunction32. Furthermore, oxidative stress triggers a cascade of events that lead to the degeneration process of dopaminergic neurons in PD51. Increased oxidative stress might also impair the convective flow and CSF-to-ISF turnover, which causes glymphatic dysfunction52. DTI-ALPS was shown to be associated with oxidative burden conditions in PD, indicating the possible role of oxidative stress in the glymphatic system29. These risk factors and pathogenic pathways possibly contribute to the impairment of the glymphatic system, which are yet to be studied in future studies.DTI-ALPS index could be regarded as a non-invasive biomarker to measure the fluid dynamics in the brain and represent the potential glymphatic function, as well as offer insight into fluidopathy in PD. To a certain extent, decreased DTI-ALPS indexes are linked to impairment of glymphatic function25 and provided better-resolution DTI images to calculate the DTI-ALPS index. We report two candidate imaging biomarkers, DTI-ALPS index and PVS, which were complementary in evaluating the glymphatic function, and thus provides a more complete assessment. There are some limitations to the current study. First, we conducted this study in a single center, and the sample size was relatively small for subgroup analysis, which needs further verification in future multi-center study with a larger sample size. Second, we speculated, based on our results, that the glymphatic dysfunction might initiate from the left hemisphere and progress to the right hemisphere in our study sample, since close to 70% of the PD patients had right-sided symptom onset. However, no differences were observed in DTI-ALPS indexes between right-onset and left-onset PD patients. Whether the left to right propagation of the glymphatic dysfunction stands and the possible presentation of developing synucleinopathy cannot be concluded from our results, and long-term prospective studies are needed. Thus, future research is warranted, which will help to build the multi-modality biomarker profile for PD.The strength of this study includes using both DTI-ALPS and PVS burden indexes to assess the glymphatic functional and structural abnormality, and to assess the asymmetry of fluidopathy in the brain using ultra-high field 7 T MRI. The 7 T MRI improved the visualization of PVSIn conclusion, this study demonstrated that the DTI-ALPS indexes were decreased in PD, which might initiate from the left hemisphere and progressively involve the right hemisphere with the disease progression. Decreased DTI-ALPS index correlated with increased PVS burden, indicating that both metrics could reflect brain glymphatic function. MRI evaluation of the diffusion aberrations near PVS potentially could be an imaging biomarker for PD disease progression in evaluating both motor and non-motor features.1. A total of 48 community volunteers without neurological or psychiatric disorders were recruited as HCs. HCs had no family history of PD or related neurodegenerative disorders. The PD patients were divided into two subgroups according to their Hoehn & Yahr stage (H&Y stage). PDa group included 40 patients whose H&Y stage was lower than or equal to 2, while the PDb group contained 36 patients with H&Y stage higher than 2. To match for age and sex for the PDa and PDb groups, 47 controls were assigned to HCa, and 31 control participants were assigned to HCb group. Some HCs overlapped between the two groups.A total of 76 PD patients were recruited from the Second Affiliated Hospital, and Sir Run Run Shaw Hospital of Zhejiang University School of Medicine. All PD patients were evaluated by movement disorders neurologists. The diagnosis was based on the International Parkinson & Movement Disorder Society (MDS) Clinical Diagnostic Criteria for PDFor clinical evaluation, PD patients were assessed using UPDRS I to IV and H&Y stage during defined off-stage (PD medications on hold for a minimum of 12 h). We applied HAM-A and HAM-D to assess anxiety and depression. The MMSE was used to evaluate cognitive function during the on-stage of patients.The study was approved by the ethics committee of the Second Affiliated Hospital of Zhejiang University School of Medicine (Ethics No. 2015-081) and the ethics committee of Sir Run Run Shaw Hospital of Zhejiang University School of Medicine (Ethics No. 20200908-30), respectively. Written informed consent was obtained from all the participants.3 × 0.7 mm3 × 0.7 mm3, TR = 5000 ms, TI1/TI2 = 900/2750 ms, TE = 2.3 ms, α1/α2 = 5°/3°, and two times generalized autocalibrating partial parallel acquisition acceleration), a T2-weighted turbo spin echo sequence , a multiband echo-planar DTI sequence , and a susceptibility-weighted imaging (SWI) sequence . Patients underwent MRI scan during defined off-stage (PD medications on hold for a minimum of 12 h).MRI images were acquired with a 7 T Magnatom research system  with prototype sequences, including a magnetization prepared with two rapid gradient echoes (MP2RAGE) sequence (voxel size: 0.7 mmhttp://www.fmrib.ox.ac.uk/fsl) to conduct processing steps, including topup correction, eddy current correction, DTIFIT, registration to standard space. Color-coded fractional anisotropy (FA) maps and diffusivity maps in directions of the x-, y-, and z-axes  were generated. DTI-ALPS method was applied to evaluate the activity of the glymphatic system29. At the level of the lateral ventricle body, the SWI image showed that the medullary veins ran perpendicular to the ventricular wall  to visualize the projection, association and subcortical fibers that run perpendicular to each other  and along the z-axis of association fibers (Dzassoc) were also extracted. The major difference for the water molecule behavior between the mean diffusivity along x-axis in projection (Dxproj) and association (Dxassoc) fibers, and mean diffusivity along the y-axis in projection fibers (Dyproj) and along the z-axis of association fibers (Dzassoc) might come from the involvement of PVS6. The DTI-ALPS index was defined as follows: ALPS index = mean /mean 6. The DTI-ALPS indexes in both hemispheres were calculated.The DTI data were processed using FSL . PVS number was assessed and PVS volume was calculated in both hemispheres. Following the manual delineating along the boundary of all identified PVS, the ITK-SNAP software automatically gave the voxel number of identified PVS, and PVS volume was calculated as the sum of the individual volume of identified PVS.PVS burden of the BG was analyzed as described previously46 in both periventricular white matter (PVWM) and deep white matter (DWM) on T2-weight MRI images. For each region of white matter, the ratings of participants were scored from 0 to 3. Furthermore, the total Fazekas score ranging from 0 to 6 was calculated by summing the two ratings.The WML were assessed according to the Fazekas rating scalet-test was applied for the comparison of continuous variables with normal distribution, Mann–Whitney test was used to compare continuous variables with non-normal distribution. Chi-squared test was used for the comparison of categorical variables. Pearson correlation analysis for normally-distributed data, whereas Spearman correlation analysis for non-normally-distributed data were performed to further evaluate the correlations between DTI-ALPS indexes and clinical characteristics, including disease duration, UPDRS, H&Y stage, LEDD37, MMSE, HAM-A, and HAM-D scores. To investigate whether the alteration of the DTI-ALPS index was consistent with PVS burden in PD, the relationship between the two metrics were analyzed. Pearson correlation coefficient (pcc) and Spearman correlation coefficient (scc) were calculated, respectively. P value <0.05 was considered statistically significant. We further performed a false discovery rate (FDR) approach for multiple testing, and FDR-adjusted p value < 0.05 indicated survival from multiple testing. ROC curve was used for the sensitivity analysis of DTI-ALPS indexes for PD patients. We also combined multiple biomarkers, including the DTI-ALPS indexes and PVS number/volume together into a single composite score, using multiple logistic regression54. Then we used the ROC curves to evaluate the quality of the composite score.Statistical analyses were conducted with GraphPad Prism . The Shapiro–Wilk normality test was used to test the normality of variable distribution. Two sample Further information on research design is available in the supplementary materialReporting Summary


The Ponderal index, a measure of a newborn’s leanness, was negatively associated with maternal urinary Pb  and Hg  (both p < 0.05). No association between maternal Cd and birth outcomes was observed. Most heavy metals showed positive associations with free cortisol and 8-OHdG. Free cortisol was identified as a mediator underlying the observed relationship between Hg and birth length or Ponderal index. This study observed adverse birth outcomes from maternal exposures to Pb and Hg. Increased free cortisol related to Hg exposure was suggested as a possible causal pathway from Hg exposure to birth outcomes such as the Ponderal index.Prenatal exposure to heavy metals is known to be associated with adverse birth outcomes and oxidative stress biomarkers. In this study, we examined whether maternal free cortisol or 8-Hydroxy-2-Deoxyguanosine (8-OHdG) could mediate associations between maternal heavy metal exposure and birth outcomes. A total of 182 healthy pregnant women were recruited. Heavy metals , free-cortisol, and 8-OHdG were analyzed in urine at delivery. Birth outcomes including birth weight, length, Ponderal index, and head circumference were measured. To examine associations of maternal urinary heavy metals with biomarkers and birth outcomes, generalized linear models were employed. Birth length was positively associated with Pb  and Hg  (both  Prenatal exposure to heavy metals is well-known to be related to adverse health outcomes of fetuses ,2,3. StuHeavy metal exposure can lead to an overproduction of reactive oxygen species (ROS) ,8,9. ROSUrinary 8-Hydroxy-2-Deoxyguanosine (8-OHdG) is commonly used as a biomarker of ROS-mediated DNA damage ,8. StudiCortisol, the primary stress hormone, is a steroid hormone produced in the adrenal gland following the activation of the hypothalamic-pituitary-adrenal (HPA) axis . It is nPreviously, we examined heavy metal exposure in pregnant women, exposure sources, and health effects on their children from the Children’s Health and Environmental Chemicals in Korea (CHECK) cohort ,24,25,26In the stage of sampling, a total of 335 mother–child pairs in the CHECK cohort were recruited using the following inclusion criteria: gestational age ≥ 37 weeks, birth weight ≥ 2500 g, no medical predisposition, and no occupational exposure history to metals. Among a total of 335 mother-child pairs in the CHECK, 182 pairs were included in the present study after excluding pairs with no matching data for a combination of heavy metals and biomarkers in urine (n = 133) or missing data of specific gravity of urine (n = 20) . This stbirth weight (g)]/[birth length (cm)]3 × 100) [A systematic questionnaire was used to obtain demographic and pregnancy-related information, including maternal age, pre-pregnancy BMI from maternal height and weight, income, smoking (active and passive) or drinking during pregnancy, delivery mode, and parity. Birth outcomes including sex, birth weight (g), birth length (cm), head circumference (cm), and gestational age at delivery were retrieved from birth records. Ponderal index, an alternative index to BMI, was calculated with the formula of [3 × 100) . A low P3 × 100) .Maternal spot urine samples were collected when they arrived at the hospital right before their delivery. All urinary samples were stored at −80 °C until analysis. Pb, Cd, and Hg, the three most frequently reported heavy metals, were analyzed. Pb and Cd were analyzed with graphite furnace atomic absorption spectrometry . Total Hg was analyzed using Mercury Analyzer . Furthermore, in the quality assurance and quality control processes for measurements of urinary metals, the accuracy and recovery ranges calculated using the spiked urine were 90–110% and 85–100%, respectively. External Quality Assessment Scheme  was also used. Detailed analytical procedures were provided elsewhere ,29. LimiUrinary 8-OHdG was measured using a competitive enzyme-linked immunosorbent away with an 8-hydroxy-2-deoxy Guanosine EIA kit  . The LODUrinary concentrations of Pb, Cd, Hg, free cortisol, and 8-OHdG were adjusted by specific gravity (SG) to compensate for urine dilution . SG has In order to investigate mediation effects of biomarkers (free-cortisol or 8-OHdG) on the association between heavy metals and birth outcomes, we conducted causal mediation analyses. Unstandardized indirect effects were computed for each of 1000 bootstrapped samples, and 95% confidence intervals were computed by determining indirect effects at the 2.5th and 97.5th percentiles. The total effect was the effect of maternal heavy metal exposure on the outcome. The indirect effect was the effect of heavy metal exposure on birth outcomes mediated through maternal biomarkers. The direct effect was the effect of maternal heavy metal exposure on the birth outcome without maternal biomarkers. The percent mediated was calculated as the value of the indirect effect divided by the sum of the indirect effect and direct effect . The conceptual model for mediation analysis in this study is illustrated in http://cran.r-project.org, accessed 15 March, 2020) was used for all statistical analyses.Sensitivity analyses were performed to evaluate the robustness of the mediation analysis by considering covariates that might be associated with heavy metal exposure and birth outcomes. In sensitivity analyses, fish and seafood intake were also considered. The intake of fish and seafood was calculated as serving/month and divided into two groups, based on the median, a high intake group or a low intake group. R version 3.6.1  concentrations. Urinary Pb was higher (p < 0.03) for mothers who consumed alcohol during pregnancy. Urinary Hg showed a trend of decrease (p < 0.05) with increasing pre-pregnancy BMI. Urinary Hg was also significantly different by household income. However, no trend was observed. Smoking during pregnancy and parity did not affect maternal heavy metal concentrations. Concentrations of heavy metals were not associated with gestational age or delivery mode .Heavy metals were detected in most urine samples, with detection rates of 91.8%, 99.5%, and 96.7% for Pb, Hg, and Cd, respectively . The medery mode . The Spep = 0.046, Free cortisol and 8-OHdG were detected in all maternal urine samples. The median (range) of free cortisol was 34.2 (5.19–178) μg/dL and that of 8-OHdG was 60.5 (29.0–601) ng/mL. Free cortisol and 8-OHdG were moderately correlated  and Cd , while 8-OHdG was positively associated with Pb , Hg , and Cd .p = 0.020) or 0.89 cm  increase in birth length, respectively  or −0.27 g/cm3  decrease in the Ponderal index, respectively  increase in birth length and a 0.32 g/cm3 decrease in the Ponderal index , as shown in results of analysis with covariates adjusted. No significant association between 8-OHdG and birth outcomes was observed. Mediation analysis was performed to understand the relationship between urinary heavy metals and birth outcomes through biomarkers. Significant mediation effects of free cortisol on birth length (the percent mediated 19.4%) and the Ponderal index (the percent mediated 30.3%) were observed in relation to Hg in maternal urine . Other tConcentrations of maternal urinary Pb, Hg, and Cd in this study were compared with previous studies . ConcentConcentrations of Hg in maternal urine samples of this study  were slightly lower than those in China   and EgypConcentrations of Cd  in this study were slightly higher compared to those in other Asian countries, including Bangladesh (median: 0.58 μg/L) , China (Positive associations of urinary free cortisol and 8-OHdG levels with heavy metals were observed in this study , consistSeveral studies have reported associations of heavy metals with increased cortisol levels ,54. CortIncreased maternal urinary Pb levels were positively associated with birth lengths of newborns. However, they showed no significant associations with birth weight a. The poA negative association was observed for maternal urinary Pb with the Ponderal index of newborns in the present study. The Ponderal index is a good measurement to identify symmetrical or asymmetrical growth status in neonates . The lowHg exposure showed a positive association with birth length but a negative association with Ponderal index in the present study. Its associations with body weight and head circumstances tended to be negative, although such associations were not statistically significant b. IncreaNo significant associations were identified between Cd and birth outcomes in this study c, while Negative associations of free cortisol levels with birth circumstance and Ponderal index were observed, whereas free cortisol showed a positive association with birth lengths of newborns . It is w8-OHdG showed no significant association with any birth outcomes in the present study . Excess Studies that examine the association between maternal exposure to Pb, Hg, and Cd and birth outcomes considering the mediation effects of 8-OHdG and free cortisol have been rarely reported. There is a cohort study suggesting that mercury-induced oxidative stress might induce adverse pregnancy outcomes, such as spontaneous abortion, preeclampsia, and lower birth weight in newborns. However, the study did not address the mediation effect of oxidative stress in the association between Hg exposure and negative birth outcomes . To the This cross-sectional study has limitations. Spot urine samples collected at delivery might not accurately reflect free cortisol or 8-OHdG levels over the course of pregnancy. Previous studies have reported that free cortisol and 8-OHdG concentrations are increased as pregnancy progresses ,84. In aAssociations between maternal urinary heavy metals including Pb, Hg, and Cd and the anthropometric outcomes of newborns were assessed in consideration of free-cortisol and 8-OHdG levels in maternal urine. Maternal Pb and Hg levels were associated with increased birth length but decreased Ponderal index. Free cortisol and 8-OHdG levels were elevated with increasing levels of heavy metals in maternal urine. Mediation analysis suggested that maternal exposure to Hg might induce adverse birth outcomes mediated through increased free cortisol. Such mediation effects were not observed for Pb exposure or 8-OHdG. Further studies are necessary to investigate potential mechanisms underlying such associations between heavy metal exposure and birth outcomes.