Within the first weeks after insertion of the second LNG-IUS, she developed a depressive syndrome and anxieties. An extensive somatic, including gynecological examination revealed no pathological findings and a mental disorder was suggested. Due to the patient´s request and the recommendation of her psychiatrist, the device was removed and led to a remission of her mental complaints up to a 6- and 12-months follow-up. Beyond the mood changes considerably affecting her quality of life, the patient raised the concerns that she has never been informed about potential ADRs on mental health and her remarks regarding the potential association between psychiatric symptoms and the LNG-IUS were considered as groundless. With this case, we strengthen previous observations regarding mood changes under LNG-IUS. Moreover, we illustrate that psychiatric symptoms may also occur as ADRs during the subsequent insertion. Thus, we emphasize that psychiatric symptoms have to be clearly communicated as ADRs to patients with LNG-IUS within a written informed consent and should be routinely examined by gynecologists.The levonorgestrel-releasing intrauterine system (LNG-IUS) is used as hormonal contraception by millions of women worldwide. It is considered as a safe device with low rates of systemic adverse drug reactions (ADRs). However, an emerging evidence suggest mood changes as ADRs. Whereas most of these studies report psychiatric ADRs after the first implantation of the LNG-IUS, it has to be considered that these may also occur after replacement, even when psychiatric symptoms were not evident at the time of the initial insertion. A potential explanation for the development of psychiatric ADRs in subsequent LNG-IUS may rely on fluctuations of sex hormones throughout the female life cycle with changing windows of vulnerabilities for developing mood disorders. Thus, the reliable contraception for women remains a continual challenge. We present the case of a 41-year-old woman that used the LNG-IUS (Mirena The levonorgestrel‐releasing intrauterine delivery system (LNG-IUS) is registered in more than 120 countries worldwide and considered as an effective and safe device for contraception for up to 5 years, and for the treatment of several gynecological conditions, e.g. menorrhagia [5]. The most frequent adverse drug reactions (ADRs) of LNG-IUS are thought to be menstrual irregularities and changes in bleeding patterns –6 arisesWe illustrate the case of a 41-year-old woman who developed a severe depressive syndrome with panic attacks and suicidal ideations after insertion of the second LNG-IUS after 5 years. These symptoms remitted after the removal of the device.®) was chosen as a long-term contraception owing to the patients` preference. Of note, there was no other gynecological indication (e.g. menorrhagia) for the insertion of the device. Except for daily tasks involving her household and taking care of her four children, no psychosocial stress factors were reported. There was no history of psychiatric disorders or general medical condition, neither in the past nor within her family. The patient reported from a regular menstrual cycle and denied current or previous pre- or perimenstrual syndromes. Moreover, there was no other medication than the LNG-IUS within the last few years. After 5 years of implantation of the first LNG-IUS and approximately 2.5 years from present, a second LNG-IUS was inserted. The removal of the first and the insertion of the new LNG-IUS occurred on the same day. Within the first weeks after the insertion of the second device, she experienced an increasing psychomotor restlessness that was pronounced in the evening and during the night. She also reported from episodes of tachycardia and experienced an irregular heartbeat. Within the next 6 months, symptoms worsened and she developed sleep disturbances, anxiety in the night, emotional lability and suicidal ideations. She also reported of somatic complaints such as shivering, hot flashes, and pelvic pain with variable intensity. Repeated medical examinations by the family doctor, including ECG, blood tests  were without any significant findings. A cardiologic examination including a long-term ECG, exercise ECG and cardiac ultrasound revealed supraventricular extrasystoles without any other pathological findings. These internal and cardiological non-findings could however, not calm the patients´ worries down and had no impact on her well-being. Her anxieties still worsened with an increased frequency of panic attacks including the fear to faint or even to die and led to an emergency call. Due to the pelvic pain, a rheumatological examination including the determination of autoantibodies  was conducted but also without any pathological findings. Several gynecological examinations led to the presumptive diagnosis of a premenopausal syndrome in the context of an early menopause. However, since the previous somatic examinations revealed no pathological findings, a mental origin was assumed and a psychiatric treatment was recommended. At this time the duration of the symptomatic has worsened over the course of 3 years. The 41-year-old housewife was married for 20 years. After the birth of her youngest child 8 years ago, an LNG-IUS  and mirtazapine (7.5 mg OPD). This medication was taken daily for 6 months. In addition, lorazepam (0.5–1 mg) was prescribed as medication on demand. With medication, she reported from a minor improvement of sleep disturbances and from a remission of panic attacks, but still, higher levels of anxiety, depressed mood, inner restlessness, rumination and emotional lability persisted. Thus, the patient consulted a psychiatrist and underwent a supportive psychotherapy for five sessions in total. At this time, psychometric subjective measures of depressive mood conducted with the Beck depression inventoryTwo weeks after removal of the LNG-IUS, psychiatric and in particular depressive symptoms improved markedly up to a BDI sum-score of 18, indicating a mild depressive syndrome, despite reduction of antidepressants. After two further weeks, the antidepressive medication was stopped. Four weeks after removal of the LNG-IUS, the patient reported that depressive symptoms and anxiety remitted completely (BDI sum-score: 6). In a 6- and 12-months follow-up, there were no clinically relevant symptoms of depression or anxiety (BDI: 2 respectively 2) observed.® was not conducted for ethical reasons. Despite the well-known discussion regarding the latency of antidepressive medication in which they achieve their full effects  drug-monitoring-center , the def effects , one mayThe recent literature support a good benefit-risk assessment for LNG-IUS as contraception with low systemic effects , 20, preIn contrast to this recommendation, a nationwide prospective cohort study in Denmark observed an increase in suicide attempts and even suicide associated to the use of LNG-IUS . NotablyThus, our observation is in line with previous studies reporting a burden of affective symptoms in LNG-IUS users, suggesting a 34-percent higher risk of depression in women using the LNG-IUS . Of noteWhereas most of these studies investigated mood effects after the first insertion of the device, it is of note that our patient developed symptoms of depression and anxiety after the implantation of the second LNG-IUS. In general, it is well known that side effects of medication occur even after the intake of years. However, considering the increasing evidence that in particular women are at higher risk to develop mood symptoms due to significant fluctuations of sex hormones over time , one mayvia ultrasound to ensure adequate contraception, it should be equally usual for gynecologists to examine the patient’s mental condition, not only after first insertion of an LNG-IUS and over time, but also in particular after replacement and insertion of the subsequent device. Moreover, this case should emphasize the inevitable necessity to clarify potential psychiatric side effects in terms of a written informed consent.Nevertheless, our case is supposed to strengthen the awareness of every health worker for potential ADRs, and in particular psychiatric symptoms. While it is quite usual to control the positioning of the LNG-IUS Since I was very satisfied with the LNG-IUS, I decided to replace the device after the recommended 5 years. Shortly after the insertion of the second LNG-IUS, I became increasingly nervous, felt an inner unrest and during the following weeks, I felt sad and had even suicidal thoughts. I consulted several medical doctors, but they found no physical cause for my complaints. My family doctor told me that I might have a depressive disorder, prescribed antidepressants and referred me to a psychiatrist. After the intake of the antidepressants, I felt somewhat relieved as the intensity of the symptoms became less, but nevertheless, I still felt sick and limited in various settings of my daily life and my quality of life heavily decreased. My psychiatrist told me that the LNG-IUS might be causative for my mental symptoms and recommended to remove it. In contrast, my gynecologist mentioned that the LNG-IUS may not have an impact on my mental status. Since may mood still further decreased and since I previously suggested an association between the LNG-IUS and my psychological complaints by myself, I repeatedly asked my gynecologist to remove the device. Within the next few weeks after the removal and although I quit antidepressive medication, I felt increasingly better, my mood was getting better and my inner restlessness remitted. I have no doubts that my mental complaints were associated to the LNG-IUS considering the strong association between the onset of my symptoms and the removal. I am aware that every medication has side effects that cannot be predicted. Nevertheless, I am still disappointed and angry since my concerns were not taken seriously and more important, I was not informed regarding potential mental symptoms before the insertion of the device, neither by verbal information, nor by a written product information.The original contributions presented in the study are included in the article/supplementary material; further inquiries can be directed to the corresponding author.Written informed consent was obtained from the individual(s) for the publication of any potentially identifiable images or data included in this article.RZ, CS-L, and HG wrote the manuscript with support from MG. CS-L was in contact with the patient. All authors contributed to the article and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Electrokinetic remediation is a useful technique for the removal of ionic contaminants in soils, sediments, sludges, and other solid porous matrixes. The efficiency of metal removal and the electricity consumption in the electrokinetic treatment of soils largely depend on electric and physicochemical conditions. This study analyzes the electrokinetic treatment of Mn contaminated kaolinite clay specimen and the influence of voltage, current intensity, moisture content, pH, and facilitating agents on metal removal and energy consumption. The objective of this study is to identify the influence of the typical variables used in electrokinetic remediation. The results showed that the operation at constant voltage or constant current intensity were equivalent in terms of metal removal and energy consumption, as long as the electric field intensity was kept low to minimize the consumption in parallel electrochemical reactions, especially the electrolysis of water. The moisture content had a significant influence on the Mn removal. Moisture content higher that 50 percent resulted in very effective Mn removal as compared with kaolinite specimens with lower moisture. The control of pH in the electrolyte solutions and the addition of facilitating agents (organic acids) enhanced the removal of Mn but increased the electric energy cost. Overall, the best conditions for Mn removal involved low to moderate electric potential difference (10 to 30 V), the use of citric acid as the facilitating agent, and the pH control in the cathode at a slightly acid pH. The electrokinetic treatment of a sludge from a water treatment plant contaminated with Mn was effective when pH control on the cathode was used. Mn and various metals  were removed with moderate electricity and acid consumption. Electrokinetic remediation is a soil decontamination technology extensively studied from the 1990s in the twentieth century. The electrokinetic treatment of contaminated soils is based on the application of a low intensity electric field directly to the soil with two main electrodes, i.e., anode and cathode. The electric field induces the mobilization of the contaminants and their transportation towards the electrodes. The contaminants are removed by two main mechanisms, electromigration (the transport of ions towards the electrode of opposite charge) and electroosmosis . Furthermore, the electrochemical reactions induced by the electric field on the electrodes has an enormous influence in the speciation of the contaminants and their transportation out of the soil. The main reactions induced by the electric field are the electrolysis of water that generates H+ ions in the anode and OH− ions in the cathode. This reaction and the pH changes induced by the H+ and OH− ions are decisive in the efficiency of electrokinetics for the removal of metals from contaminated soils [In 2009, Reddy and Cameselle ,4. There−, NO3−, etc.) [The electrokinetic remediation has been studied for the removal of heavy metals and toxic elements  ,16, othe−, etc.) ,18, and −, etc.) ,20 such −, etc.) , the ele−, etc.) . The eff−, etc.) . As it w−, etc.)  and the −, etc.)  to keep −, etc.) .This work focuses on the assessment of the influence of the electric field strength and its mode of application to the soil in the electrokinetic removal of heavy metals. In order to understand the influence of the electric parameters, a simple model soil composed of kaolinite clay contaminated with manganese sulfate was selected for this work. This model soil simplified the geochemistry of the soil for the identification of the influence of the electric field on physicochemical parameters , and on the transportation of the contaminant metals. In addition, the electric energy consumption and the treatment time were considered in the study as fundamental parameters for the design of practical applications and large scale. The Mn results and conditions obtained in the tests with model soil samples are used for real samples of an industrial sludge contaminated with Mn. The electrokinetic tests with model samples could help in defining the operating conditions of the real samples. However, the electrokinetic remediation of real samples requires further study and optimization since the geochemistry and aging of contaminants is different than the model samples and they will affect the results . This stThe electrokinetic cell used in all the tests was designed in our research group based on a previous design by Pamucku et al.  from the2/g. Mineralogy analysis by X-ray diffraction showed the presence of 85% kaolinite clay, 14% mica, and 1% quartz. The particle size was selected based on the availability of kaolin commercial products from the company CAVISA. The smallest size (3 μm) was selected to simulate the behavior of a low permeability clayey soil. Kaolinite was selected since it shows much lower buffering capacity and cation-exchange capacity as compared with other clay minerals. The model soil specimen was prepared mixing 100 mL of concentrated manganese sulfate solution with 120 g of dry kaolinite. The mixture stood overnight (about 24 h) before it was used in the electrokinetic tests. The final mixture had a moisture content of 45% and a concentration of 5 g Mn/kg of dry kaolinite.The model soil specimen used in all the tests was composed of kaolin contaminated with manganese sulfate. Kaolin, supplied by the company CAVISA , has an average particle size of 3 μm and a specific surface area of 13.5 mThe spiked kaolinite was introduced in the central tube of the electrokinetic cell using the consolidation unit. This consolidation unit created homogeneous and saturated soil specimens for the electrokinetic tests. During the consolidation process, a fraction of water and manganese was lost. The final Mn concentration in the kaolinite specimen ranged from 3.6 to 3.8 g Mn/kg of dry soil and the moisture content ranged from 37% to 40%. The initial pH of the contaminated kaolinite was pH = 4 to 4.5.A sludge from a water treatment plant in an industrial area was used to test the capability of electrokinetic remediation to remove metals from real solid matrixes. The sludge was contaminated mainly with Mn, although it contained significant amounts of other metals .The contaminated kaolinite with Mn or the industrial sludge was introduced in the central tube of the electrokinetic cell and it was compacted in the consolidation unit. Then, the electrode chambers were attached to the central tube and filled with deionized (DI) water. The main electrodes were connected to the direct current (DC) power supply and the electrode chambers were connected to the flow panel. A constant DC electric potential (10 to 30 V) or electric current intensity (1.2 mA) was applied through the main electrodes for a maximum of 7 days (except some specific tests which duration is specified in the results section). Periodic readings of electric current intensity and electric potential among the main and auxiliary electrodes were recorded. The electroosmotic flow was measured by volume variations in the flow panel. The pH of the electrode solutions was measured along the testing time and registered. At the end of the test, the electrode solutions were collected and analyzed for pH and metal concentration. The kaolinite specimen  was extruded from the central tube and divided into 5 equal portions or sections, namely S1 to S5 from anode to cathode. A sample of the initial kaolinite specimen  and the samples from the 5 final portions were analyzed for metal concentration, pH, and moisture content.Moisture content in kaolinite or sludge samples was determined by dry weight at 105 °C until constant weight. The pH in the solid samples was determined mixing 1 g of solid and 2.5 mL of DI water for 1 h, and then the pH was measured in the supernatant fluid. The Mn in the liquid and solid samples was determined by inductively coupled plasma optical emission spectrometry (ICP-OES) with a piece of equipment Optima 4300 DV from Perkin Elmer . The samples were firstly digested following the USEPA method 3010A  or method 3050B .The electromigration of Mn in contaminated kaolinite specimen was firstly tested at a constant electric potential gradient of 30 V. This value was selected based on previous electrokinetic tests with metal contaminated kaolinite ,28 where+ ions, and the reduction of water in the cathode yielded OH− ions. Thus, the anode was acidified and the cathode was alkalinized  hydroxide, Mn(OH)2, showed a white-yellowish color, but this compound easily transformed into a mixture of oxides and hydroxides with the general formula MnO(OH) with a characteristic brown color [The pH and Mn distribution in kaolinite specimen at the end of the experiment can be explained based on the pH changes induced by the electrolysis of water in the electrode chambers. The oxidation of water in the anode yielded Halinized b. The acalinized c, desorbwn color . Thus, t2+, and the precipitation of manganese in Section S5 explain the electric current intensity profile and the distribution of the potential gradient in the system  produced at the cathode. The amount of electricity consumed in the electrolysis of water is compared with the total charge that passed through the electrokinetic cell in terms of mol of electrons  and the cathode chamber.The operation at a constant electric potential or electric current intensity are the two alternative modes for any electrochemical test. The operation at a constant electric potential difference between anode and cathode implies that the driving force for the transportation of ions is constant. Thus, the total flux of ions, i.e., the total charge through the system, varies along the treatment time depending on the availability of ions, in other words, the electric conductivity of the system. The operation at constant electric current intensity means that the flow of charge, i.e., the flow of ions, is constant, and the voltage gradient varies depending on the availability of ions to transport the charge between anode and cathode.2 . This is an intermediate value of the current intensity registered in the previous test at 30 V. This value was selected to be able to compare the results of this test at a constant current intensity and the previous test at a constant 30 V. The results of Mn removal and electric power consumption are shown in 2+ ions started to accumulate in Section S5. The lower removal of Mn in Sections S1 to S4 was due to the lower driving force  developed in the system because of the use of such a low electric current intensity. This was also the reason for the lower electric power consumption.The test at a constant electric current intensity was carried out at 1.5 A/mOverall, the response of the electrokinetic system seemed to be similar at a constant current intensity or a constant electric potential. It is important to keep the electric field intensity low to decrease the electricity spent in the electrolysis of water. Low voltage or current intensity would result in low energy consumption, but the removal of metal would require more time.The profile of the electric current intensity in 2).As can be seen in 2+) from the anode to cathode, but at the same time, an increase in voltage also increases the total electric power consumption due to the major extension of parallel reactions, i.e., the electrolysis of water. As it can be seen in The electric potential difference used in the electrokinetic tests could have a significant influence on the removal of Mn and electric power consumption. Voltage increases the driving force for the transportation of cations  depending on the soil pH and the charge of the predominant species. The literature results proved the efficiency of the complexing agents removing metals from real soil samples, but the results were very affected by the pH.An alternative method to overcome the premature precipitation of Mn in the last section of kaolinite S5 a is the 2+ ions and the organic acid. In the pH range used in these tests, the formation of anionic complexes is not favored, the most probable complex species are neutral or cationic [In this study, the removal of Mn from model kaolinite specimens was tested using five organic acids with complexing capacity, i.e., acetic acid, EDTA, citric acid, tartaric acid, and oxalic acid . The orgcationic ,36. Thuscationic ,38, citrThe electrokinetic treatment was applied to a sludge from a water treatment plant in an industrial area. The plant treated the runoff water from a mining area by neutralization and precipitation of metals in solution and particles in suspension. The sludge contained a significant amount of metals and carbonates with a pH of 7.60. In this condition, it seemed that the best electrokinetic treatment should involve the depolarization of the reduction of water in the cathode  and favor the acid front generated in the anode (Test EK2 in +/kg of sludge) to depolarize the cathodic reduction of water must be considered in the design of an electrokinetic treatment at a large scale.As can be seen in The application of a constant electric potential of 30 V to the kaolinite specimen for 7 days resulted in a removal of Mn as high as 83% in Sections S1–S4 (4/5 of the kaolinite specimen). Mn mainly accumulated in the last section of kaolinite (69%) and cathode (14%).The operation at constant voltage or constant current intensity yielded similar results in terms of Mn removal and electric power consumption.The operation at low voltage (10 V) for longer time (40 d) increase the removal of Mn and decreased the electric power consumption as compared with the test at 30 V for 7 days.The increase of the moisture content up to 60% resulted in over 90% Mn removal in the whole kaolinite specimen.The pH control in the cathode resulted in the complete removal of Mn from the kaolinite, but a significant increment in the electric consumption was observed. The increase in the electric consumption could be restrained using pH close to the neutrality, avoiding very acid pH in the cathode.The use of organic acids as facilitating agents improved the removal of Mn. The test with citric acid showed complete removal due to the formation of Mn complexes and the enhancement of the electroosmotic flow.The electric power consumption was very affected by the extension of parallel reactions, i.e., the electrolysis of water. Any modification of the electrokinetic system  that increased the electric conductivity of the system resulted in an increase of electric power consumption.The electrokinetic treatment of an industrial sludge contaminated with Mn and other metals was only effective with the depolarization of the electrolysis of water in the cathode with sulfuric acid. This acid was selected because it does not interact with the target metal Mn.The electrokinetic removal of Mn from model kaolinite samples and real industrial sludge is effective under the following selected operating conditions:Overall, the results of the electrokinetic test in model and real samples suggested the feasibility of the electrokinetic treatment for the removal of Mn from contaminated matrices.


Men play indispensable role in health and wellbeing of mothers and children. Their level of involvement with regards to birth preparedness and complication readiness is understudied. This study was therefore carried out to assess male involvement and associated factors in birth preparedness and complication readiness in Debre Berhan town, North East Ethiopia.st - 30th, 2014 in Debre Berhan town among 806 study participants. Cluster sampling technique was employed to select study participants. Data were collected using a structured and pre-tested questionnaire by face-to-face interview technique. Bivariate and multivariate analyses were performed to check associations and control confounding.Community based cross-sectional study was conducted from July 1The study showed that male involvement in birth preparedness and complication readiness found to be 51.4%. Educational status of tertiary level , having 1 or 2 children  and 3 or 4 , knowledge of danger signs , knowledge of birth preparedness and complication readiness  and attitude  were significantly associated with male involvement in birth preparedness and complication readiness.Male involvement in birth preparedness and complication readiness found to be low in study area. Education, number of children, knowledge on danger signs, knowledge on birth preparedness and complication readiness and attitude were factors associated with male involvement. Creating awareness on danger signs of pregnancy, birth preparedness and complication readiness both at community and institutional level were recommended in order to increase male involvement in birth preparedness and complication readiness. Maternal mortality has continued to be a major challenge to health systems worldwide , 2. Althst – 30th, 2014. The town is located in the North Shoa zone of the Amhara Region; 130km from the capital city of Ethiopia, Addis Ababa in the North East direction and at 795km from the capital city of the region (Bahir Dar). Administratively, Debre Berhan town is divided into 9 kebeles and an estimated population size of 65,214 [α/2 = 1.96 . Hence, the calculated sample size was = 384. Since the sampling technique was cluster, design effect of 2 was considered, for possible none response rate 10% and the final sample size was 845. The data was collected by using pretested, structured and interviewer administered questionnaires. The questionnaires were administered by five BSc midwives. They had two days of training including the supervisors for the purposes of data collection. The data collection process was closely supervised by one field supervisor who has Bsc degree in public health and the principal investigator. In this study male involvement in birth preparedness and complication readiness was measured based on seven indicators. Thus, those respondents who respond to 3 indicators were considered as involved in birth preparedness and complication readiness. Adequate knowledge of danger signs was considered when the respondents respond to greater than or equal to five key danger signs during pregnancy, delivery and post-natal period, out of ten danger signs. The respondents who knew greater than or equal to two components of birth preparedness and complication readiness out of four components were considered as they had adequate knowledge of birth preparedness and complication readiness. Data was checked for completeness and consistency, coded, entered and cleaned with Epi-info version 7 and exported to SPSS  version 16 for analysis. The results were presented in the form of tables, and text using frequencies and summary statistics such as mean, standard deviation, and percentage to describe the study population in relation to relevant variables. The data were analyzed using logistic regression to determine the effect of various factors on the outcome variable and to control confounding. Crude and adjusted odds ratios with their 95% confidence intervals were calculated. Ethical clearance was obtained from Ethical Review Committee of University of Gondar, Department of Midwifery. A formal letter of cooperation was written to Debre Berhan town health department and each selected kebele administrations. Voluntary verbal consent was obtained from each study participant.A community based cross sectional study design was conducted in Debre Berhan town from July 1Socio-demographic characteristics of study participants: a total of 803 of husbands whose wives were pregnant or had children less than one year old were included in the study with a 95% response rate. The median age of the respondents was 33 with inter quartile range of nine. Majority of the respondents were Amhara (88.2%) by ethnicity. Regarding their educational status, majority had attended tertiary and above educational level (44.7%). More than half of respondents were not employed (58.5%) and the median income of the respondents was 2800 ETB (Ethiopian Birr) per month with inter quartile range of 1501ETB. Almost all of the respondents had monogamous marriage (99.9%) ( (99.9%) .Male involvement in birth preparedness and complication readiness: the study found out that 51.4%  of males involved in BP/CR. From the components, majority of husbands had adequate knowledge on BP/CR (64.4%) followed by who had adequate knowledge on danger signs (53.4%); and small number of husbands planned to have SBA (14.3%). Husbands whose wives were pregnant better met the components than husbands who had children less than one year old. For instance, with regard to knowledge on danger sign, 36% of husbands whose wives were pregnant, but 16.7% of husbands who had children less than one year old had adequate knowledge on danger sign  more likely to be involved in BP/CR than those who can’t read and write. With respect to the number of children, the study finding showed that those who had 3 or 4 children were 5 times  more likely to be involved in BP/CR than those who had no child. Husbands who had adequate knowledge on danger sign were about 8 times more likely to be involved than those who had no adequate knowledge on danger signs of pregnancy, childbirth and post natal period ). Similarly, those husbands who had adequate knowledge on BP/CR were about 12 times  more likely to be involved in BP/CR than those who had no adequate knowledge. Husbands who had good attitude towards male involvement in BP/CR were about 2 times more likely to be involved in BP/CR than those who had poor attitude   .The current study found out that the male involvement in BP/CR was 51.4%. This result is higher than the result found in Tigray region Enderta woreda which was 26.90% .This difHusbands who attended tertiary education were more likely to be involved in BP/CR than those who can’t read and write. This is consistent with the studies done in different African countries and other countries such as India, Guatemala and Nepal -19. ThisKnowledge of danger sign is also one of the factors that have an effect on male involvement. Those husbands who had adequate knowledge on danger sign were more likely to be involved than those who had no adequate knowledge on danger signs of pregnancy, childbirth and post-natal period. This result is similar with the findings from the researches done in Harari public health institutions, Eastern Ethiopia and other countries such as Nigeria and Nepal , 23, 24.Knowledge of husbands on BP/CR has significant effect on their involvement in BP/CR. Those husbands who had adequate knowledge on BP/CR were more likely to be involved in BP/CR than those who had no adequate knowledge. This result is similar with research done in India . This miMale involvement in birth preparedness and complication readiness was found to be low. Socio demographic factors (education and number of children); individual related factors  were significantly associated with male involvement in birth preparedness and complication readiness. Creating awareness on danger signs of pregnancy, birth preparedness and complication readiness both at community and institutional level were recommended in order to increase male involvement in birth preparedness and complication readiness.Magnitude of male involvement in BPRC is low;Known factors for high male involvement BPRC was having good attitude towards birth preparedness and complication readiness.Attending tertiary level of education;Having good knowledge on danger sign and birth preparedness and complication readiness.


Contemporary outcome data of preterm infants are essential to commission, evaluate and improve healthcare resources and outcomes while also assisting professionals and families in counselling and decision making. We analysed trends in clinical practice, morbidity, and mortality of extremely preterm infants over 10 years in South Wales, UK. This population-based study included live born infants < 28 weeks of gestation in tertiary neonatal units between 01/01/2007 and 31/12/2016. Patient characteristics, clinical practices, mortality, and morbidity were studied until death or discharge home. Temporal trends were examined by adjusted multivariable logistic regression models and expressed as adjusted odds ratios (aOR) with 95% confidence intervals (95% CI). A sensitivity analysis was conducted after excluding infants born at < 24 weeks of gestation. In this population, overall mortality for infants after live birth was 28.2% (267/948). The odds of mortality  and admission to the neonatal unit  significantly decreased over time. Non-invasive ventilation support during stabilisation at birth increased significantly  with corresponding decrease in mechanical ventilation at birth  and following admission (0.80 [0.68–0.96]). Medical treatment for patent ductus arteriosus significantly decreased over the study period . The incidence of major neonatal morbidities remained stable, except for a reduction in late-onset sepsis . Gestation and centre of birth were significant independent factors for several outcomes. The results from our sensitivity analysis were compatible with our main results with the notable exception of death after admission to NICU . There were significant improvements in survival and reduction of late-onset sepsis of extreme preterm infants in South Wales between 2007 and 2016. The sensitivity analysis suggests that some of the temporal changes observed were driven by improved outcomes in the most preterm of infants. Clinical practices related to respiratory support have changed but significant variations in clinical practices and outcomes between centres remain unexplained. The adoption of regional evidence-based clinical guidelines is likely to improve outcomes and reduce variation. Over the last few decades, survival rates of extremely preterm infants have steadily increased6 due to improvements in antenatal care and neonatal intensive care management. Despite improved survival, the incidence of morbidities unique to this population has not shown statistically significant reduction over the last decades6.Although extreme preterm infants born at a gestational age of < 28 weeks are a small proportion of all births7. Trend analyses of outcomes linked to clinical practices are crucial in identifying interventions that can lead to improved outcomes. Such analysis can also help to direct appropriate resources to support and promote specific areas of clinical practice, perpetuating a culture of continuous improvement and driving the establishment of research networks and quality improvement programmes worldwide8.Medical teams taking care of extremely preterm infants need up-to-date outcome data to be able to counsel families and aid decision making10. Data from England and Wales show that 7% of all live births are preterm . Of this 11% are very preterm  and 5% are extreme preterm 11. Extreme preterm infants are at higher risk of severe morbidities and mortality3.Compared to other European countries, the UK has higher rates of extreme and very preterm birthsThe three tertiary neonatal units in South Wales have been caring for extreme preterm infants for decades, and clinical care practices have undergone major changes over time. A comprehensive analysis of outcomes to guide local counselling of parents and to identify areas of improvement have not been conducted. We hypothesised that mortality, major morbidities, and care practices in this vulnerable group of infants would show statistically significant improvement over time, as reported in the literature. The objective of this study was to report the temporal trends of key outcomes . This neonatal care for the population is centralised and served by three tertiary neonatal intensive care units (NICUs) that offer a full range of medical care for preterm infants; one of these units also serves as a regional centre for neonatal surgery and other subspecialty services. Extreme preterm infants needing critical care were either born at one of the tertiary centres or transferred into these centres soon after birth. Besides, six local neonatal units provided limited high-dependency and special care for the more mature preterm infants closer to home either from birth or following repatriation from the tertiary units.South Wales is home to 75% of the population of Wales, with England as its eastern border and mid-Wales forming the northern border. The estimated birth rate in South Wales is around 25,000 per year13.The study cohort included all infants born alive at a gestational age of < 28 weeks between 1st January 2007 and 31st December 2016 at one of the three tertiary neonatal units in South Wales (inborn infants). This included live born infants who died in the delivery room following unsuccessful resuscitation. Infants born at < 28 weeks at any of the local neonatal units and transferred into one of the three tertiary units were also included in the cohort (outborn infants). No exclusion criteria were applied; however, extreme preterm infants who were born alive at local neonatal units but died before transfer to a tertiary unit were not included in the cohort. As a variable known to affect outcomes, statistical analysis accounted for the differences in the place of birthVON) for all infants born at < 30 weeks of gestation or with birth weight < 1500 g. Data for the VON database was collected and verified by a designated senior clinician in each unit, ensuring consistency and high-quality. This study included prospectively collected anonymised data from all three tertiary neonatal units over a 10-year study period for infants < 28 weeks of gestation at birth.From 2007, all three tertiary neonatal units contribute data for benchmarking purposes to the Vermont Oxford Network  and are described in detail in Supplementary Table Raw data included maternal and neonatal characteristics, resuscitation practices, early clinical management, and clinical outcomes including death and major morbidities. The data was collected for each infant for their first admission to the NICU until death or transfer to another centre or discharge home from the tertiary unit. Standard VON definitions were used for major morbidities .Sex (binary variable).Place of birth, i.e. inborn or outborn (binary variable).Hospital of care .The following unmodifiable co-variates were used for adjusted analyses:Mortality after live birth and after admission to NICU.14 OR periventricular leukomalacia.Severe cranial ultrasound (CUSS) abnormalities, defined as a composite of severe intraventricular haemorrhage (IVH) grade III–IVSurgery for retinopathy of prematurity (ROP).Bronchopulmonary dysplasia (BPD) at 36 weeks, defined as a requirement for mechanical ventilation, non-invasive ventilation, or supplemental oxygen at 36-weeks post-menstrual age.Medical treatment for a patent ductus arteriosus (PDA) with indomethacin or ibuprofen.15.Necrotising enterocolitis (NEC), defined as Bell stage II–IIISepsis, defined as a culture-positive episode with bacterial or fungal organisms.The co-primary outcomes included the following major outcomes:As death was a competing variable for all the major morbidity outcomes, a composite of death with each of the major morbidities was also analysed.Secondary outcomes included key changes in respiratory practice including the use of surfactant, use of non-invasive respiratory support and use of mechanical ventilation.Statistical analysis was undertaken in SPSS version 25 . Missing or unavailable data were appropriately coded and excluded from analysis at all stages. These are presented in Supplementary Table In the first block, the dependent variable was modelled against the year of birth as an independent (continuous) variable and presented as unadjusted odds ratio (OR).16 , the centre of care12  and outborn status17  and presented as adjusted OR. Individual centres were not compared with each other; summary results testing centre as a factor are presented if reached statistical significance.In the second block, the regression analyses were adjusted for confounders by including gestation at birth , genderThe third block was used to derive the most parsimonious model for each major outcome. All independent variables  from block 2 were included in the model along with the clinically relevant interaction variables gestation at birth*year of birth, gestation at birth*gender and centre of care*outborn-status. A stepwise variable selection procedure (forward selection) was specified in which variables were sequentially entered into the model. The first variable considered for entry into the equation was the one with the largest positive or negative correlation with the dependent variable (outcome). This variable was entered into the equation only if it satisfied the criterion for entry . If the first variable was entered, the independent variable not in the equation that had the largest partial correlation was considered next. The procedure stopped when there were no variables that met the entry criterion.Trends were analysed by deriving multivariable logistic regression models in blocks with the outcome-of-interest  as the dependent variable.Due to the possibility of significant variation in the clinical care of infants born < 24 weeks of gestation, a sensitivity analysis of the co-primary and secondary outcomes was undertaken only for infants with a birth gestation between 24 and 27 weeks using the same methods as above.A p-value of < 0.05 was considered statistically significant. All uses of the word “significant” in the results section refer to statistical significance only.guidance tool for ethics approval. There was also no direct involvement of patients or their parents/guardians as none were identifiable. A data-sharing agreement was signed and approved by the NHS Research and Development Department at each site. Categorical variables with n < 5 have been suppressed in tables to prevent accidental identification of individuals and sites.As this analysis was conducted on non-identifiable information, which was previously collected routinely for clinical purposes, it was excluded from specific ethical approval as per general guidance on the NHS Health Research Authority +0 and 23+6 weeks, 314 (33.1%) between 24+0 and 25+6 weeks and 534 (56.3%) between 26+0 and 27+6 weeks. The birth weight ranged from 310 to 1500 g  and the head circumference at birth (812 available) ranged from 18 to 37.4 cm . A slight male preponderance was recorded i.e. 54.7% . Almost a quarter of the infants  were multiple births, but the number of multiple pregnancies decreased over the study period . A major birth defect was reported in 5.3% (50/948) of the study population and this trend did not change significantly over time (p = 0.19). Detailed results on infant characteristics, maternal characteristics, resuscitation at delivery and respiratory support of infants after resuscitation are presented in Table In the 10 years, 948 infants born before 28 weeks of gestation were cared for in one of the three tertiary NICUs in South Wales; 741 (78.2%) were inborn. Median gestation of this population was 26.3 weeks (IQR 2.3 weeks). Of them, 100 (10.6%) were born between 22Figure Of the 898 infants admitted to a neonatal unit, 875 (97.4%) had a cranial ultrasound scan (CUSS) during their stay. Of these scans, 137 (15.7%) reported a severe intraventricular haemorrhage (IVH). Periventricular leukomalacia (PVL) was an uncommon finding  and did not change significantly over time (p = 0.08). The composite outcome of severe CUSS changes (severe IVH OR PVL) was noted in 151 infants (17.2%) decreased significantly with increasing gestation  but did not change over time . The composite of death OR severe CUSS changes decreased with increasing gestation  and over the study period . The proportion of missing data was small, primarily contributed by infants who died in the delivery room or within 12-h who did not have a CUSS performed.Incidence of surgery for ROP remained stable over the study period , although the composite of death OR ROP surgery reduced marginally . Odds for both outcomes reduced with increasing gestation (p < 0.01 for both), and there was a significant centre effect (p < 0.01 for both).Of the 401 infants for whom data was available, 290 infants (72.3% of available data) developed bronchopulmonary dysplasia (BPD) after live birth. The incidence of BPD did not change significantly over the study period  but the odds of developing BPD was significantly lower at the higher gestational ages . Accounting for deaths, the odds of the composite outcome death OR BPD  did not change over the study period , although this decreased with increasing gestation . However, there was a significant proportion of missing data (57.7% for BPD and 30.5% for death OR BPD).The odds of medical treatment for diagnosed PDA with either indomethacin or ibuprofen decreased significantly over time  and was less likely with increasing gestation .Of the 898 infants who were admitted to a neonatal unit, 106 (11.8%) were reported to have necrotising enterocolitis (NEC) and 63 (7%) had surgery for NEC, suspected NEC, or bowel perforation. The odds of NEC were stable over the study period  but significantly decreased with increasing gestational age . Outborn infants had significantly higher odds for developing NEC during the study period . The centre of care had a significant effect on this outcome (p < 0.01), most likely due to cases being concentrated in one surgical centre.During the study period, 340 infants developed sepsis; of these, 33 were early-onset (before 72-h of life) and the rest were late-onset (beyond 72-h of life). The odds of late-onset sepsis (LOS) reduced over time , with increasing gestation  and in boys . Missing data for sepsis was due to the early death of infants and the data was complete for the composite of death OR sepsis.The odds of using non-invasive respiratory support during resuscitation significantly increased  while mechanical ventilation during  or after  resuscitation significantly decreased over the study period. Use of surfactant decreased marginally over the study period . All these outcomes were dependent on gestation, with the use of non-invasive ventilation increasing with gestation (p < 0.01) and mechanical ventilation decreasing with gestation (p < 0.01).After excluding the 100 infants who were born at < 24 weeks of gestation, a sensitivity analysis was conducted for the co-primary and secondary outcomes. Point estimates  with a corresponding decline in the odds of endotracheal intubation (14%). More mature infants had higher odds of being intubated at birth, possibly secondary to the emerging use of INSURE technique to administer early surfactant to facilitate rapid extubation and before the increasing popularity of using LISA techniques. Mechanical ventilation on conventional modes at any time during stay significantly reduced over the study period vindicating our hypothesis that infants were spending less time on mechanical ventilation at any time over the study period, although there was a modest but significant increase in the use of high-frequency ventilation. Use of the most recent mode of neonatal respiratory support—high flow nasal cannula therapy—significantly increased over the study period21.In keeping with current evidence22. The results from our sensitivity analysis, after excluding infants < 24 weeks of gestation, were compatible with our main results with the notable exception of death after admission to NICU. This possibly suggests that some of the temporal changes in mortality which were observed in the whole cohort may have been driven by the improved outcome in the most preterm infants born at < 24 weeks of gestation.Cohort studies on extreme preterm gestation suggest that the approach to infants < 24 weeks of gestation is variable; some of them may not receive active resuscitation or care23 but half that of a comparable population in the US3. Similarly, severe IVH and severe ROP was higher in our population than in other international and European studies19. Although the reasons for these differences in outcomes are unclear, it could be related to better survival in our population. We found a higher incidence of IVH but a lower incidence of LOS in boys compared to girls; this contrasts with the pattern reported before and warrants further research. Although the treatment for PDA remains subjective, our results are in keeping with reported trends from the USA24. It is important to point out that the incidence of BPD has reportedly been on the rise in other comparable populations from the USA3 and UK18. While our stable trend for most neonatal morbidities may be reassuring to some, one may argue that the lack of improvement may reflect the lag in translating improving practices from research studies20 into clinical practice19. This slow pace is not uncommon in trend analysis of population outcome8.The incidence of the major neonatal morbidities  remained largely static over the study period except for LOS. Although the decreasing trend was reflective of the overall UK trend, the incidence of LOS reported in our population was higher than published UK data25, exploring the cause for this difference was beyond the scope of this paper and likely to be multifactorial. The adoption of evidence-based network and national guidelines should significantly reduce some of the variations observed but must be balanced with the need for tailoring individualised care and clinician’s autonomy for wise professional judgment.We reported significant variations in multiple areas of clinical practice as well as outcomes between participating centres. These included practices related to resuscitation, respiratory support, and management of PDA as well as outcomes such as IVH, ROP and sepsis. Differences in rates of surgery could be explained by the fact that only one of the centres offered neonatal surgical care. While variation between centres is well known in neonatal practiceLike other retrospective studies from clinical databases, we had to account for missing data in several data fields (Supplementary Table 26, and local population differences.Our study has several limitations. While the large majority of the extreme preterm infants born alive in South Wales were included in our analysis, infants who died in the delivery room at local neonatal units before transfer to the tertiary units could not be captured in this data set. Although our study spanned 10 years, the number of infants in each gestation week was still insufficient to undertake a meaningful subgroup analysis. We have adjusted for known confounders, but other unmeasured factors may also have had an impact on the outcomes presented. It was beyond the scope of our study to explore the link between care practices and outcomes; this is planned for a later stage. We have used ‘centre’ as a simple adjusting variable which hides further complexity including differences in care practices, the intensity of work27. We have restricted our analysis to the extreme preterm population who are at the highest risk for morbidity and mortality after birth. Our population was well defined due to the model of centralised neonatal care as part of a managed clinical network in South Wales. The inception point for outcomes was ‘live birth’ which effectively included all infants born with signs of life, thus reducing selection bias. The outcome definitions are embedded in the internationally accepted VON criteria, and additional outcomes have been defined clearly before analysis including the timing of assessment where relevant. Information bias was likely to be minimal considering that data input into the VON database was undertaken in each unit by senior clinicians with substantial experience and VON data collection and interpretation of outcome definitions. Our analysis is detailed, longitudinal, adjusted for common confounders, and reports uncertainty ranges for all outcomes.However, our study has several strengths which need highlighting. We have followed recommendations for reporting outcomes of extreme preterm infants28. Research into more accurate prediction30 and prevention28 of preterm delivery should become the focus of healthcare and research funding bodies worldwide to enable further improvements in outcomes of preterm infants.Extreme preterm infants in South Wales remain vulnerable to multiple major morbidities and mortality after birth. While opportunities for improvement in care and outcomes remain and are the topics of current research studies, major changes can only be expected from a significant reduction in the preterm population being born at this extreme range of gestation. Currently, no effective treatment options are available to significantly delay or prevent premature birthIn summary, we have reported detailed outcomes of an extreme preterm population who were born alive over 10 years in Wales. This is the first such report from Wales, and our data will help counsel parents and plan further improvements in South Wales.Supplementary Information.


There is ample evidence of associations between short birth interval and adverse maternal and child health outcomes, including infant and maternal mortality. Short birth interval is more common among women in low- and middle-income countries. Identifying actionable aspects of short birth interval is necessary to address the problem. To our knowledge, this is the first systematic review to systematize evidence on risk factors for short birth interval in low- and middle-income countries.A systematic mixed studies review searched PubMed, Embase, LILACS, and Popline databases for empirical studies on the topic. We included documents in English, Spanish, French, Italian, and Portuguese, without date restriction. Two independent reviewers screened the articles and extracted the data. We used the Mixed Methods Appraisal Tool to conduct a quality appraisal of the included studies. To accommodate variable definition of factors and outcomes, we present only a narrative synthesis of the findings.Forty-three of an initial 2802 documents met inclusion criteria, 30 of them observational studies and 14 published after 2010. Twenty-one studies came from Africa, 18 from Asia, and four from Latin America. Thirty-two reported quantitative studies , 10 qualitative studies, and one a mixed-methods study. Studies most commonly explored education and age of the mother, previous pregnancy outcome, breastfeeding, contraception, socioeconomic level, parity, and sex of the preceding child. For most factors, studies reported both positive and negative associations with short birth interval. Shorter breastfeeding and female sex of the previous child were the only factors consistently associated with short birth interval. The quantitative and qualitative studies reported largely non-overlapping results.Promotion of breastfeeding could help to reduce short birth interval and has many other benefits. Addressing the preference for a male child is complex and a longer-term challenge. Future quantitative research could examine associations between birth interval and factors reported in qualitative studies, use longitudinal and experimental designs, ensure consistency in outcome and exposure definitions, and include Latin American countries.CRD42018117654.Prospectively registered on PROSPERO  under registration number  Adequately spaced births allow women to recover from previous pregnancies, but both too short or too long intervals lead to adverse maternal, perinatal, neonatal, and child health outcomes . The WorA meta-analysis published in 2006 reported that short < 18 months) and long > 59 months) intervals between two consecutive pregnancies were associated with preterm birth, low birth weight, and being small for gestational age 8 months , and oth9 months A systematic review published in 2012 suggested potential mechanisms for the adverse consequences of short birth interval . These iOne estimate suggests that around 2 million of the 11 million deaths per year of children under 5 years old could be prevented by avoiding birth intervals of less than 2 years . InternaShort birth interval is more common among women in low- and middle-income countries, where an estimated 17% of married women of reproductive age are reported to have unmet needs for family planning . For exaSeveral systematic reviews have examined the consequences of birth interval for maternal and child health outcomes , 3, 5. AWe registered the protocol prospectively on PROSPERO  under registration number CRD42018117654 . Our resUsing key terms specific to each database, Boolean operators, and truncators, we developed the search strategy with the collaboration of an experienced librarian from McGill University. We included PubMed, Embase, LILACS, and Popline, which is a database specialized in maternal and reproductive health containing grey literature . Our seaWe used the following inclusion criteria: (i) experimental or observational studies reporting a measure of association , qualitative studies, and mixed methods studies; (ii) studies in English, Spanish, French, Italian, and Portuguese; (iii) studies conducted in low- or middle-income countries according to the World Bank classification ; (iv) exp-values (without a measure of magnitude of association), studies from high-income countries, studies using “birth spacing” as a synonym for contraception use, and studies exploring factors associated with the interval between marriage and first birth.We excluded theoretical publications, quantitative studies reporting only proportions or Using the open-source systematic review web application Rayyan , two indThe research team collectively designed the data extraction form based on the variables that would answer the research question, in an iterative process with regular meetings to discuss and update the form. JP and UA independently piloted the data extraction form on 5% of the studies to determine its appropriateness. They then extracted data from the included studies and conducted a cross-check review to verify the quality and accuracy of the extracted data.We extracted the following data when available: basic study information ; population and setting ; study methods ; and results .We planned to conduct a quantitative synthesis only if the included studies were sufficiently homogeneous and of adequate quality. As definition of factors was heterogeneous and study quality was variable, we performed a narrative synthesis of the quantitative findings and an inductive thematic analysis  on the rTo assess the quality of the included studies, we used the 2018 Mixed Methods Appraisal Tool (MMAT), a quality appraisal instrument for systematic reviews including qualitative, quantitative, and mixed methods studies . ResearcFrom an initial 2802 documents identified by our search, 43 studies remained after screening and assessment , education of the mother (six), and contraception use (five). Five studies defined short birth interval as < 24 months, while two studies used < 33 months and < 36 months, respectively . For all factors but length of breastfeeding (three studies) and sex of the previous child (two studies), researchers reported mixed results. Table Seven studies found an association between a short birth interval and a younger age of the mother. Among these publications, two studies explored age at last delivery, one study explored the age of the mother at the moment of the study, and the remaining studies did not specify a definition of age of the mother. Two studies reported an association between a short birth interval and older age of the mother (one study explored age at first pregnancy).Three studies found an association between short birth interval with shorter duration of breastfeeding. One study found more short birth interval with no breastfeeding as opposed to exclusive or mixed breastfeeding.Three studies reported an association between a short birth interval and no contraception use, while one publication reported the contrary. The latter study used data from Demographic and Health Surveys from nine sub-Saharan African countries between 1991 and 2001.Five studies reported more short birth interval with less education, but one study reported more short birth interval with more education. Two studies reported an association between short birth interval and less parity, while one study reported the contrary.Two studies reported more short birth interval with lower income, while one study reported the contrary. Similarly, a study reported an association between short birth interval and fewer assets. One study reported more short birth interval for people living in rural areas compared with people living in urban settings.Three studies reported an association between short birth interval and an adverse outcome of a previous pregnancy, but one study found longer birth interval after a previous abortion. One study found more short birth interval after a singleton birth compared with after a multiple birth. One study found more short birth interval when the place of delivery of the index child was not a health facility. Two studies found more short birth interval when the sex of the previous child was female.An experimental study found more short birth interval with maternal and newborn health care only, compared with integrated post-partum family planning and maternal and newborn health care. Four studies reported other factors associated with birth intervals such as marital status of the mother, occupation of husband, place of residence, year, and religion Table .After adjusting for confounders, six studies reported non-significant associations between different factors and short birth interval. The studies explored age of the mother at the moment of the study (four publications), education of the mother (three), contraception method, marital status of the mother, occupation of the mother, and place of residence (two publications each category). Other factors examined included contraception use, age at last delivery, age of the woman’s mother at first pregnancy, type of breastfeeding, occupation of the husband, parity of the mother, delivery place of the previous pregnancy, planned previous pregnancy, and income level (one publication each category).Interpretation of time-to-event data is challenging since the nature of association may change over time. For example, more education can lead to longer birth intervals in the first two births, but it may have the opposite effect for subsequent births. We have summarized the authors’ overall conclusions for each study , previous pregnancy outcome (seven), age of the mother (six), breastfeeding (four), and socioeconomic level (three). All the studies reported mixed results for all factors except for breastfeeding, for which four studies found more short birth interval with shorter breastfeeding in all the pregnancies.Three studies reported shorter birth intervals with a younger age of the mother, while one study reported the contrary. Two studies that looked at age at marriage and birth interval reported mixed results, and one study found shorter birth intervals with younger age at first birth.Four studies found shorter birth intervals with lower education of the mother, while three studies found the contrary, and two other studies found mixed results. Five studies found shorter birth intervals with adverse outcomes in any previous pregnancy, while one study found the opposite. Two studies found mixed results.One study reported a shorter birth interval with roundworm (Ascaris lumbricoides) infection but extended interbirth intervals with hookworm infection. Another study found longer birth intervals among women with HIV infection.One study reported shorter birth intervals among married couples, while another study found shorter birth intervals with shorter duration of the marriage. Regarding socioeconomic level, a study found shorter birth intervals with lower income, another study found a shorter birth interval with rural residence, and a third study found mixed results (place of residence).Six studies found shorter birth intervals with less parity, postpartum amenorrhea less than 6 months, when the previous child is a girl, not using modern contraception, when religion is Catholic, farmer as a profession, and less female decision-making autonomy. An intervention study found longer birth intervals with early short-term infant supplementation. A study found mixed results on birth interval for media exposure and employment status of the women.We identified nine themes among the findings of the qualitative studies. The leading theme was local concepts and practices (14 codes), followed by modernization (eight), communication (six), religion (four), breastfeeding, health concerns, knowledge and attitudes, miscellaneous (two codes each theme), and autonomy (one).agbon, described as a “female body odor after parturition” that is to be respected before restarting sexual activity; apa, which requires avoiding a conflictive situation between children if spaced too closely; and ratat, a traditional period of abstinence. Researchers also reported the influence of local medicine people , traditional medicine, use of amulets, talismans, and cords, coitus interruptus, polygamy, and social taboo on the length of birth interval.According to the participants of the studies, the observance of local traditions prevents short birth interval. Examples include Modernization was another concept reported by researchers. The participants of the studies explained that modernization introduces social changes influencing birth intervals such as loss of culture and traditions, men staying at home more often, availability of health services, education, and food, and changes in religion and beliefs. Availability of health services and infrastructure, and family planning education prolong birth interval, while the rest of the factors related to modernization promote short birth interval.According to the participants of the qualitative studies, birth interval was also affected by communication between couples and families, the influence of local media and the society, and observation of other parents. Concerns for maternal and child health were mentioned as tending to increase birth interval, as was knowledge about contraception methods.Some participants mentioned that breastfeeding prolongs birth interval, while other participants questioned its effectiveness as a means of preventing an early next pregnancy. Finally, Catholicism and Hinduism were considered to shorten birth spacing, and women’s autonomy, drought, and war were also mentioned to affect the timing of births. Table We decided not to conduct a meta-analysis. The included quantitative studies were very heterogeneous in their definitions of exposures and outcomes, and most used a cross-sectional design with variable approaches to dealing with potential confounders.The quality of most documents ranked as medium (23 documents), followed by high (16 documents), and low (four documents). For information about the quality assessment please see Additional file Our systematic review shows two factors consistently associated with short birth interval: shorter breastfeeding and a female previous child. Younger age of the mother, less education of the mother, a negative outcome of the previous pregnancy, and lower socioeconomic status were often associated with short birth interval, although some studies reported the opposite. The quantitative studies examined a limited number of factors that could be easily included in a questionnaire.A contraceptive effect of breastfeeding has long been recognised to prolong birth interval. Breastfeeding causes lactational amenorrhea because the suckling stimulus downregulates hypothalamic gonadotropin-releasing hormone secretion and the production of luteinizing hormone .In 1988, an international group of scientists met at Bellagio, Italy , with thDespite current efforts to promote breastfeeding worldwide, early weaning is still common and few children receive exclusive breastfeeding by the age of 6 months . A recenPreference for a son is widespread in North Africa, East and South Asia, and the Middle East . In thesStudies reported mixed results for associations with a negative outcome of the previous pregnancy, age of the mother, education, and socioeconomic level, although with a preponderance of findings in one direction in each case. Researchers conducting the studies in this review often reported an association between adverse outcome of the previous pregnancy, including abortion, pregnancy loss, and multiple birth, and short birth interval. The author of one of the included studies  suggesteMost studies found more short birth interval with a younger age of the mother. Younger women are generally more fertile and more sexually active; very young women are likely to be economically disadvantaged, and may have less access to and use of modern contraception, which can explain this association –36, thouMost studies reported that less-educated women have more short birth interval. Hailu  explainsMost studies in our review found more short birth interval among economically underprivileged women. Hailu  suggestsSimilarly, most studies reported expected associations between longer birth intervals with modern contraception use. One cross-sectional study based on DHS surveys reported opposite results  and the Some of the studies in this review failed to find significant associations between age of the mother, education, contraception method, marital status, occupation, and place of residence, and short birth interval. Together with the fact that some studies showed associations in the opposite direction to the prevalent findings, this suggests that associations with factors such as age and education of the mother are not universal and may be highly dependent on the particular context.Qualitative studies explored the perceived effects of local concepts and practices not considered in quantitative studies. For example, in Burkina Faso  and KenyQualitative studies considered other factors that were not examined in quantitative studies. Participants in qualitative studies mentioned that communication between couples and families, the influence of local media and the society, and observation of other parents could influence birth intervals. None of the included quantitative studies attempted to explore associations between these factors and birth interval.Quantitative studies reported a consistent association between longer breastfeeding and longer birth interval. However, participants in one qualitative study questioned the effectiveness of breastfeeding as a means to prolong birth interval, presumably based on their own experiences . The WHOStrengths of our study include a broad search strategy guided by a librarian, inclusion of five languages, no year limit, and inclusion of quantitative, qualitative and mixed-methods research, and grey literature.We chose not to conduct a meta-analysis due to considerable heterogeneity of study design, study quality, population characteristics, and outcome and exposure definitions. We noted the concerns of Egger and co-authors  that (i)As with all systematic reviews, our results and conclusions are limited by the quality of the original studies. Almost all the included quantitative studies were observational, with concerns about residual confounding even after multivariate analyses , and temThe variability of the definition of short birth interval  limits the conclusions of our study. The current WHO recommendation of an optimal birth interval .We found two factors consistently associated with short birth interval: shorter breastfeeding duration and a female previous child. Promotion of breastfeeding could help to reduce short birth interval, and has many other benefits . AddressThe quantitative and qualitative studies yielded different and complementary findings. This highlights the potential value of mixed-methods research. Quantitative researchers should look for ways to investigate factors such as local knowledge and practices, modernization, and communication. Future research should use longitudinal and experimental designs, aim for consistency in outcome and exposure definitions, and include Latin American countries.Additional file 1. PRISMA 2009 Checklist. Filled PRISMA checklist showing 27 items and the page reporting each item.Additional file 2. Search strategy. Key terms specific to each database, Boolean operators, and truncators.Additional file 3. Full references of the records included in our study. List showing the full references of the documents included in our study.Additional file 4. Studies reporting HR. Table showing the findings of studies reporting HR.Additional file 5. Quality assessment of the studies. Table showing the quality appraisal scores (two reviewers and average).


Bifidobacterium longum, Bifidobacterium adolescentis, Bacteroides dorei, Bacteroides ovatus, Roseburia hominis, Faecalibacterium prausnitzii, and Eubacterium hallii. Finally, in a novel model to assess host–microbe interactions  co-culture) fermented cRG-I increased barrier integrity while decreasing markers for inflammation. In conclusion, by using novel in vitro models, cRG-I was identified as a promising prebiotic candidate to proceed to clinical studies.Modulation of the gut microbiome as a means to improve human health has recently gained increasing interest. In this study, it was investigated whether cRG-I, a carrot-derived pectic polysaccharide, enriched in rhamnogalacturonan-I (RG-I) classifies as a potential prebiotic ingredient using novel in vitro models. First, digestion methods involving α-amylase/brush border enzymes demonstrated the non-digestibility of cRG-I by host-derived enzymes versus digestible  and non-digestible controls (inulin). Then, a recently developed short-term (48 h) colonic incubation strategy was applied and revealed that cRG-I fermentation increased levels of health-promoting short-chain fatty acids  and lactate comparable but not identical to the reference prebiotic inulin. Upon upgrading this fermentation model by inclusion of a simulated mucosal environment while applying quantitative 16S-targeted Illumina sequencing, cRG-I was additionally shown to specifically stimulate operational taxonomic units (OTUs) related to health-associated species such as  Firmicutes, Bacteroidetes, Actinobacteria, Proteobacteria, Fusobacteria, and Verrucomicrobia phyla [®; Mucosal Simulator of the Human Intestinal Microbial Ecosystem) was recently developed as a complementary in vitro tool [Clostridium cluster XIVa members, correlating with in vivo findings from biopsies [The colon contains a vast number of bacteria that largely impact human health. Next to preventing pathogen colonization through secretion of antimicrobial agents ,2,3, theia phyla . Despitetro tool . In thisbiopsies ,16,17,18Modulation of the human gut microbiome as a route to improve human health has gained a lot of interest over recent years. Prebiotics are defined as non-digestible food ingredients that selectively stimulate health-promoting bacteria . A key fBifidobacteriaceae, prebiotic potential has often been related to an increase of this family. There is however increasing understanding that prebiotics can be fermented by a wider range of gut microbes. Inulin can e.g., also be rapidly fermented by health-promoting Bacteroidaceae members, such as Bacteroides caccae [Fructans, such as fructooligosaccharides (FOS) and inulin are considered to be “gold standard” prebiotics, with human clinical trials supporting their beneficial effect in acute and chronic diseases such as obesity and type 2 diabetes (T2D), allergy, inflammatory bowel disease (IBD), Traveler’s diarrhea and constipation (an overview is given in ). As mans caccae ,23. Currs caccae . Given ts caccae .Gut microbial modulation is linked to human health with the concept of a “leaky gut” having gained attention, not only in the context of inflammatory bowel disease, but also in a wider range of psychological and metabolic disorders . IncreasTherefore, the present study investigated whether a carrot-derived RG-I enriched extract (cRG-I) classifies as a potential prebiotic ingredient using a combination of novel in vitro models . First, ® HP, 100% inulin, 0% sweetness level, average DP ≥ 23) was generously provided by Beneo GmbH .The two carrot RG-I preparations used in this study , 2.0 g/L Yeast Extract, 2.0 g/L pepton , 1.0 g/L mucin , 0.5 g/L L-cystein and 2.0 mL/L Tween80 . When mucin-coated carriers were added to the reactors during Test 3, 1.0 g/L mucin was omitted from the nutritional medium. Five mucin-coated carriers were added per reactor after being prepared according to Van den Abbeele et al. (2013) [Short-term colonic incubations were performed as described in . Briefly. (2013) . Test prSamples were collected upon 0 h, 6 h, 24 h, and 48 h of incubation from each colon reactor. Gas production was measure with a pressure meter  and pH measurements were performed with a Senseline pH meter F410 . Total SCFA were determined as the sum of acetate, propionate, butyrate and branched-chain fatty acids  levels, and were measured as described previously . LactateT difference were omitted. The qPCRs were performed as described previously for the following groups: Lactobacillus spp. [Bifidobacterium spp. and Eubacterium rectale/Clostridium coccoides [Akkermansia muciniphila [Bacteroidetes [Enterobacteriaceae [Faecalibacterium prausnitzii [Roseburia and Eubacterium hallii [After 48 h of incubation, samples from both lumen and mucus were collected for analysis of the microbial community composition through quantitative polymerase chain reaction (qPCR) and 16S-targeted Illumina sequencing. DNA was isolated as described in  from eitlus spp. , Bifidoboccoides , Akkermainiphila , Bacteroroidetes , Enteroberiaceae , Faecaliusnitzii , Roseburm hallii . In addim hallii  to obtaim hallii . As descm hallii ,47, the m hallii  and (5) TM . In brief, blood was collected and diluted  in DPBS without Ca/Mg . Then, LymphoprepTM solution was added and samples were centrifuged at 1027× g for 20′ at room temperature (RT) to separate the mononuclear cells (MNCs) and red blood cells by density-gradient centrifugation. MNCs were collected and washed 3 times with ice-cold DPBS . Aliquots of PBMCs were frozen in liquid nitrogen. Baseline IL-8 levels were determined by enzyme-linked immunosorbent assay (ELISA)  to eliminate donors with high basal cytokine levels. For experiments, Caco-2 cells were seeded on 24-well semipermeable inserts (0.4 μm pore size) at a density of 1 × 105 cells/insert and cultured for 14 days until a functional cell monolayer with a transepithelial electrical resistance (TEER) of more than 300 Ωcm2 was obtained. PBMCs, stimulated with 2.5 µg/mL pokeweed mitogen (PWM) were added to the basolateral chamber at a concentration of 1 × 106 cells/well. PBMCs without PWM stimulation were included as negative control. At the same time, colonic suspensions (filter-sterilized (0.22 µm) and diluted 1/5 (v/v) in culture medium) or 5 mM sodium butyrate (NaB)  were added to the Caco-2 cells at the apical side. Caco-2 cells were also treated with medium as negative control. Cells were incubated for 48 h at 37 °C in a humidified atmosphere of air/CO2 . TEER was measured at start (0 h timepoint) and after 48 h of incubation using a Millicell ERS2 Voltohmmeter . All 48 h values were normalized to their own 0 h value after subtraction of the empty insert value and are presented as percentage of initial value. In addition, basolateral supernatant was collected after 48 h of incubation for cytokine analysis. Human IFN-γ, IL-17A, IL-21, IL-22, IL-4, and IL-9 levels were determined by Luminex® multiplex  and IL-10 levels were measured by ELISA , according to the manufacturers’ instructions. All experiments were performed in triplicate.Caco-2 cells  were cultured in Dulbecco’s Modified Eagle Medium (DMEM) containing glucose and glutamine  and supplemented with HEPES (4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid) and 20% (v/v) heat-inactivated (HI) fetal bovine serum (FBS) . PBMCs were isolated from buffy coats of healthy donors  using Lymphoprepp < 0.05, 2 signs = p < 0.01, 3 signs = p < 0.001 and 4 signs = p < 0.0001. Statistical analysis was performed with the GraphPad Prism software . To evaluate differences in microbial community composition at family and OTU level between blank and treatment incubations (Test 3), a Student’s t-test was performed (Excel Software). Differences were found significant if p < 0.05. To evaluate differences between PWM+ and PWM- or NaB in the Caco-2/PBMC co-culture assay (Test 4), an ordinary one-way ANOVA with Dunnett’s multiple comparisons test was performed; while differences between blank and treatment incubations were assessed with an ordinary one-way ANOVA with Tukey’s multiple comparisons test. Statistically significant differences are presented by (*). (*) = p < 0.05, (**) = p < 0.01, (***) = p < 0.001 and (****) = p < 0.0001. Statistical analysis was performed with the GraphPad Prism software .To evaluate differences in microbial metabolites (tests 2 and 3) and microbial community composition at phylum level between blank and treatment incubations (Test 3), a two-way analysis of variance (ANOVA) with Dunnett’s multiple comparisons test was performed. Statistically significant differences between the blank and treatments are presented by ,  or . 1 sign = One of the characteristics of a prebiotic is its non-digestibility by host enzymes upon passage along the upper GIT . First, Short-term colonic incubations were performed to investigate the potential prebiotic effect of cRG-I on microbial activity, including inulin as a positive control. A first indication of cRG-I fermentation by the gut microbiota followed from the significant pH decrease and enhanced gas production during the first 6 h of incubation (Δ0–6 h) versus the blank A,B. ThesUpon measuring lactate and SCFA production, acids responsible for aforementioned pH changes were elucidated. During the 0 h to 6 h time interval, cRG-I, but not inulin, significantly increased the production of total SCFA C which rAn in-depth characterization of the effects of cRG-I on microbial metabolic activity A–H was pcRG-I+LMWC exerted similar effects on microbial activity versus cRG-I with some minor differences. These included a more profound pH decrease for cRG-I+LMWC during the first 6 h of incubation due to enhanced acetate and lactate levels. Furthermore, acetate and propionate production between 6–24 h was less strongly increased with cRG-I+LMWC compared to cRG-I.Overall, while no major differences were observed between incubations with or without mucosal environment, inclusion of mucus beads led to a tendency to higher gas production, with higher butyrate levels for the blank incubation, suggesting colonization of butyrate-producing bacteria on the mucin-coated carriers.The data at phylum level are presented both as proportional A and absActinobacteria, Proteobacteria and Verrucomicrobia were enriched in the lumen (® model [Firmicutes enrichment was due to a marked enrichment in Clostridiaceae cluster I and Lachnospiraceae (Clostridium butyricum), OTU10 (related to Clostridium tertium), OTU8 (related to Clostridium paraputrificum), OTU12 (related to Ruminococcus torques), OTU3 (related to Roseburia hominis) and OTU19 (related to Ruminococcus lactaris)   . Furthera levels A,B. At fpies/mL) .Actinobacteria, Bacteroidetes and Proteobacteria compared to the blank  and Bifidobacterium adolescentis (OTU21) , B. plebeius (OTU6), B. xylanisolvens (OTU18) and especially B. dorei (OTU2). As a remark, a decrease in abundances of OTUs related to B. caccae (OTU13), B. fragilis (OTU11) and B. uniformis (OTU17) upon cRG-I and cRG-I+LMWC treatment was noted, but these decreases were less profound compared to observed increases in other OTUs. The increased abundance in Proteobacteria related to an enrichment of Desulfovibrionaceae and Enterobacteriaceae, which was most pronounced upon dosing of cRG-I+LMWC. Finally, the luminal increase in Firmicutes with cRG-I and cRG-I+LMWC was due to increased levels of Erysipelotrichaceae, Peptostreptococcaceae, Streptococcacae, Ruminococcaceae, and Veillonellaceae. The enrichment in Streptococcacae was most pronounced upon dosing cRG-I+LMWC and linked to an increase in OTU26 (related to Streptococcus aginosus). In contrast, the increase in Veillonellaceae was more pronounced upon dosing of cRG-I and was attributed to an increase in OTU9 . Within the Ruminococcaceae, two OTUs related to Faecalibacterium prausnitzii  increased upon cRG-I, while only OTU5 was stimulated by cRG-I+LMWC and this to a lesser extent.With respect to treatment effects in the lumen, cRG-I and cRG-I+LMWC both increased the absolute levels of he blank B. Althou (OTU21) , thus alreatment . FurtherFirmicutes, while decreasing Bacteroidetes levels (Lachnospiraceae (Roseburia hominis) that increased from 9.8% in the blank control incubations to 64.3% and 74.2%, respectively ; (ii) luminal Bacteroidetes; (iii) luminal Faecalibacterium prausnitzii; (iv) mucosal Roseburia ; while not affecting Akkermansia muciniphila. In addition, both products, but mostly cRG-I+LMWC, increased colonization of Eubacterium hallii in the simulated mucus layer.To confirm several of the above-mentioned observations obtained through 16S-targeted Illumina sequencing, qPCRs on specific bacterial groups of interest were performed . This coAs cRG-I fermentation enriched several health-associated species and increased production of SCFA, it might exert favorable effects at the host level. To address this question, a Caco-2/PBMC co-culture model was developed. Inflammation-induced barrier disruption was obtained upon 48 h co-culturing of Caco-2 cells with PWM-activated PBMCs and measured as a significant decrease in TEER of the Caco-2 monolayers . In addiOverall, some minor effects of the incorporation of a mucosal compartment were observed as both blank and treatment colonic suspensions slightly increased TEER; while decreased the secretion of all tested cytokines compared to luminal suspensions .® model [Firmicutes members specifically colonized the mucosal environment . Furthermore, inclusion of the mucosal environment did not alter the luminal microbiota (except for a minor enrichment in Akkermansiaceae), nor did it alter the treatment effects towards luminal microbiota activity and community composition. Overall, including the mucin beads in the current colonic simulations allowed to maintain a higher diversity, thus allowing observation of more complete treatment effects of cRG-I. Finally, application of a model combining gut epithelial and immune cells allowed to point out the gut protective effect of cRG-I fermentation-derived metabolites against an inflammatory stressor (Test 4).In the current study, novel in vitro models and analytical techniques were implemented to investigate whether cRG-I classifies as a potential prebiotic ingredient. First, upon exposure to α-amylase and brush border enzymes (Test 1), unlike the positive controls , cRG-I (like the negative control inulin) was not digested to any of the simple sugars measured. This suggests that cRG-I can be considered to be a polysaccharide that likely escapes upper GIT digestion in vivo, thus fulfilling the definition of a prebiotic ingredient that should reach the colon where it could be fermented by the gut microbiota. Secondly, a recently described short-term colonic incubation strategy  was upgr® model , a wide In a first series of short-term colonic incubations (Test 2), cRG-I was found to display prebiotic potential comparable but not identical to that of inulin in modulating microbial activity , as follBifidobacterium longum (OTU7) and Bifidobacterium adolescentis (OTU21). Bifidobacteriaceae are indeed key acetate and lactate producers [Bifidobacterium strains have been associated with beneficial effects on the host health and some strains are widely used as probiotics. A first mechanism by which Bifidobacteriaceae contribute to health is by indirectly promoting butyrate production via cross-feeding mechanisms involving for instance Faecalibacterium prausnitzii and Eubacterium hallii [B. longum strain has e.g., been shown to exert protective effects in a DSS-induced colitis model in mice by reducing inflammation and enhancing the intestinal epithelial barrier [B. longum strain reduced chronic mucosal inflammation in ulcerative colitis (UC) patients in a double-blinded, randomized-controlled clinical trial [B. adolescentis strain protected mice from DSS-induced colitis by increasing IL-10 levels, up-regulation of regulatory T-cells (Treg) and decreasing IL-17A positive T-cells [Upon fermentation in both short-term colonic incubations (Tests 2/3), cRG-I stimulated acetate, and lactate production which was accompanied with a strong reduction in pH. At community level, this correlated with increases in OTUs related to roducers ,60. Althm hallii ,61,62, tm hallii . In term barrier . Moreoveal trial . Likewis T-cells . The strPrevotellaceae and Bacteroidaceae due to the stimulation of a wide spectrum of Bacteroidaceae OTUs related to e.g., B. ovatus, B. plebeius, B. xylanisolvens and especially B. dorei. Indeed, Bacteriodetes spp. are known primary fiber degraders that are capable of producing propionate [Bacteroidaceae members have also been related to particular health benefits. As an example, in a mouse model of atherosclerosis, B. dorei reduced plaque inflammation and decreased intestinal epithelial permeability and systemic endotoxemia [B. ovatus reduced mucosal inflammation and stimulated epithelial proliferation and mucin production in a DSS-induced colitis model in mice [Veillonellaceae that was solely attributed to an increase in OTU9 . Interestingly, D. succinatiphilus is a succinate-converting, propionate-producing species [Bacteroidetes spp. are known succinate producers [Bacteroidaceae members increased upon cRG-I treatment, with OTUs relating to B. caccae and B. fragilis decreasing in abundance upon cRG-I treatment. These species are considered to be opportunistic pathogens carrying virulence factors: enterotoxigenic B. fragilis strains secrete the B. fragilis toxin (BFT) [B. caccae contains the ompW gene [Next to acetate and lactate, fermentation of cRG-I increased the production of propionate, which correlated with a stimulation of opionate . In the opionate ,8. Furthopionate . Finallyopionate . Besidesotoxemia . Further in mice . Another species  and as mroducers , its incin (BFT)  and B. cmpW gene . BesidesLachnospiraceae, containing several butyrate-producing species, in the mucosal compartment. The increase in Lachnospiraceae was related to a strong stimulation of an OTU related to Roseburia hominis, which has been associated with beneficial effects on barrier function and immune regulation in the gut [R. hominis showed protective effects in a DSS-model for colitis in mice by reducing pro-inflammatory cytokine expression. Moreover, increased Treg levels were observed in both germ-free and conventional mice fed a supplement containing live R. hominis. Also, a significant reduction of R. hominis and F. prausnitzii was observed in UC and Crohn’s disease (CD) patients [F. prausnitzii and Eubacterium hallii. Like R. hominis, F. prausnitzii was shown to exert anti-inflammatory potential by increasing IL-10 and promoting Treg differentiation in mice [E. hallii improved insulin sensitivity in a mouse model for diabetes [® study could further elucidate the potential impact of cRG-I on butyrate production.In contrast to propionate, butyrate was not significantly increased upon dosing of cRG-I. However, cRG-I did strongly stimulate  the gut . Indeed,patients ,75. Inte in mice . On the diabetes . Of notereg/TH17 axis; favoring down-regulation of excessive inflammation as IL-10 is necessary for Treg functions [reg functions has been reported in IBD and other extraintestinal autoimmune disorders [The stimulation of several health-related microbial species and the concomitant increase in health-promoting metabolites suggested that cRG-I may display interesting host beneficial properties in terms of intestinal barrier protection and reduction of inflammation. This hypothesis was confirmed using a Caco-2/PBMC co-culture model in which treatment with fermented cRG-I significantly increased the TEER, indicative for the protective effects of fermentation-derived cRG-I metabolites on inflammation-induced intestinal permeability. To further strengthen this observation, it would be interesting to perform in-depth analysis of the expression and localization of tight junction proteins including occludin, ZO-1, and claudins upon cRG-I treatment in this system. Furthermore, fermented cRG-I metabolites decreased the secretion of the pro-inflammatory cytokines IL-17A, IL-4, and IL-9, while increasing the secretion of IL-22 and of the anti-inflammatory IL-10. This is suggestive of a possible immunoregulatory effect of cRG-I metabolites on the Tunctions . Th17 ceisorders . Furtherisorders . Finallyisorders ,81. In aStreptococcacae OTU related to Streptococcus aginosus. Also, Enterobacteriaceae levels were higher upon cRG-I+LMWC treatment. Like Streptococcaceae, Enterobacteriaceae are expert fermenters of simple sugars [Coriobacteriaceae (OTU15 related to Collinsella aerofaciens) and Desulfovibrionaceae tended to be higher in incubations with cRG-I+LMWC. This suggested a potential co-existence of e.g., C. aerofaciens and Desulfovibrio piger, based on the fact that the main products of C. aerofaciens fermentation  serve as substrates for D. piger, which is the main sulfate-reducing bacterial species in the human gut microbiome [Finally, a modified formulation of cRG-I was tested which contained small size sugars, i.e., cRG-I+LMWC. Interestingly, the short-term colonic incubation strategy applied in this study was highly sensitive to pick up differences in closely related product compositions. For instance, the presence of these simple sugars resulted in a stronger initial pH decrease upon dosing of cRG-I+LMWC, related to an initial increased production of lactate, which could be linked to a stronger enrichment in a e sugars , specificrobiome . This mi® models with different fecal sample donors and in clinical trials to confirm the beneficial effect of cRG-I on the microbiota and its impact on human health.In conclusion, the implementation of novel in vitro models simulating the human colonic environment coupled to cell-based assays mimicking the host gut barrier, allowed to establish the prebiotic potential of cRG-I. This novel fiber is not digested by host enzymes characteristic of the upper gastro-intestinal tract but rapidly fermented by the human colonic microbiota leading to selective stimulation of the growth and activity of intestinal bacterial species associated with human health. cRG-I displays unique properties as it was fermented more rapidly than inulin leading to production of SCFA, especially acetate and propionate, and less gas than inulin. It increases the abundance of several bacterial species reputed for their “anti-inflammatory” profile. In line with this, the metabolites resulting from cRG-I fermentation exhibited a protective effect in an in vitro model of inflamed gut barrier. Overall, the data obtained during this study support future research to further investigate this novel prebiotic candidate in long-term SHIME