RIGINAL

Background: Severe acute malnutrition (SAM) is a state of deficiency that manifests itself in two forms: edematous and non-edematous. There have been significant advances in the clinical management of this disease state, but to date, data on nutritional recovery outcomes are scarce. Aims: We aimed to compare nutritional recovery in edematous and non-edematous malnourished children. Subjects and Methods: This is a prospective observational study of 98 children (3 – 60 months) with SAM (25 edematous and 73 non-edematous) receiving nutritional rehabilitation, according to the WHO guidelines, during a short hospitalization (SH) of 21 days. Results: Complete biochemical and nutritional recovery was completed in both edematous and non-edematous groups as indicated by normalized serum levels of retinol-binding protein, transthyretin, albumin, transferrin, fibronectin and hemoglobin, as well as C-reactive protein and acid alpha glycoprotein. Biochemical and nutritional recovery was faster in edematous children compared to non-edematous, as indicated by the significant difference (p < 0.001) in the rate of recovery of albumin, transthyretin, and retinol-binding protein. At discharge, despite full biochemical nutritional recovery there remains a severe stunting in both groups with the persistence of moderate wasting in the non-edematous group. Conclusions: Complete biochemical and nutritional recovery can be achieved after short hospitalization for nutritional rehabilitation and an early discharge of children cured is possible for edematous children but not convincing in non-edematous ones, as they remained moderately malnourished.


Introduction
Severe acute malnutrition [SAM] remains a major health concern 1 . SAM classically presents as one of two phenotypically distinct forms: edematous SAM (ESAM) and non-edematous SAM (NESAM) or marasmus 2 . In Morocco, malnutrition is qualified as a public health problem at the community level 3,4 as well as at the hospital level 5 . Considerable progress has been made in the prevention and treatment of SAM over the past 50 years. But the reviews demonstrate that the mortality rate in hospitals is approximately 20 to 30 percent in non-edematous cases and up to 60 percent in edematous ones during the past 40 years [6][7][8] . These unacceptably high mortality rates have prompted the World Health Organization [WHO] to recommend the use of guidelines for the treatment of SAM 9,10 . The goal of the WHO guideline 9,10 is to achieve better nutritional recovery by promoting the best possible treatment and shortening hospitalization in both edematous and non-edematous forms of SAM. The literature review showed that there is heterogeneity in the time of recovery from SAM, it varied from 11 days 11 to 59 days 12 . The © 2023 The Author(s). This is an open-access article. This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third-party material in this article are included in the article's Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder.
To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/. average length of stay in therapeutic centers recommended by the WHO 2 to treat SAM is 30 days, but in developing countries, most malnourished patients come from poor family backgrounds and are highly sought after. Besides, SAM is widespread in underdeveloped countries, 1 which have limited resources to deal with this public health problem. Therefore, the use of a short inpatient nutrition rehabilitation stay is advantageous because more children can be treated in a given time period at less cost 13 . As well, to have an impact at the population level, management protocols must take into consideration the realities of socioeconomic conditions, balancing the potentially conflicting demands and ethics of clinical medicine with those of public health. It is therefore essential to proactively treat complicated SAM with very intensive medical and nutritional treatments aimed at rehabilitating the child during a short hospitalization 14 . In this context, we aimed to study the pattern of nutritional recovery in children with edematous and non-edematous malnutrition after a short 21day hospitalization for nutritional rehabilitation.

Patients
According to the recommendations of the Ministry of Health, children diagnosed with moderate or severe malnutrition should be hospitalized to treat their nutritional disorders. However, in our study, every child diagnosed as severely malnourished was agreed to be hospitalized as part of standard health care. Children were enrolled in the study if they were aged 3 to 60 months and admitted as inpatients because of SAM. Children were not included in the study if their birth weight was less than 2500 g, if they had received a blood transfusion (hemoglobin concentration less than 40 g/L), if they had been referred to other departments, or if their parents had refused to participate. In addition, in our study, the children's mothers were asked to stay with their children by providing breastfeeding and frequent feeding, especially at night. All the mothers participated in the preparation of meals for their children with the help of the dietician as part of improving their nutritional knowledge. The children's mothers received nutritional education to prepare for the integration of their family environment with the aim of preventing relapses and ensuring the continuation of the child's development after discharge.

Sample size and data analysis
In order to determine the number of participants needed, we calculated the required sample size in advance using the statistical program G*Power software 3.1.9.7. In the present study, we planned to examine the difference in mean (independent means) between two groups using Student's t-test. However, we entered the required variables into the G*Power software to estimate the sample size. These variables were defined as follows: the effect size (d) was set to 0.8, alpha error probability to 0.05, power (1-β error probability) to 0.95, and the allocation ratio of N1/N2=0.45. After computation, the first group consisted of 57 children, which we increased to 73 children (non-edematous group) and the second group was composed of 25 children (edematous group). Thus, a total of 98 participants was obtained.

Study design and nutritional rehabilitation
This is an observational prospective study conducted in the pediatric unit, Ibn Khatib Hospital, Fes, Morocco. The study was designed to compare the pattern of nutritional recovery in edematous and non-edematous children after a short hospital stay (SHS) of 21 days for nutritional rehabilitation. The children hospitalized with SAM were followed-up by the head physicians of the pediatric department, the pediatricians, the dietician, as well as the nurses involved in child health care at the pediatric ward, Ibn Khatib, Fes, following the treatment recommended by the World Health Organization 9,10 . Data on children, mothers, and families were collected in a questionnaire during admission interviews. The ninety-eight children enrolled in the study were divided into two groups based on Wellcome's (1970) classification 15 : non-edematous (73 children) and edematous (25 children). During the follow-up period of nutritional rehabilitation, the healthcare team examined the children and monitored their nutritional and health status, with a progressive approach according to the standard WHO protocol 9,10 , which divides their treatment into seven stages through two phases: stabilization and rehabilitation.
Upon admission, the diagnosis was performed by pediatricians with the purpose to detect conditions that might put in danger the vital prognosis of the child. Nutrition rehabilitation started within the first 24 hours after admission. During the initial phase (3 to 9 days) the children were placed under medical and nutritional treatment. Pediatricians treated and prevented hypoglycemia, hypothermia, and electrolyte imbalance. Children with acute diarrhea (more than three liquid stools/day) were maintained on saline oral hydration solution for severely malnourished children (ReSoMal). Broad-spectrum antibiotic treatment (Gentamicin 7.5 mg/kg, Ampicillin 50 mg/kg oral, and Amoxycillin 15 mg/kg) was directly initiated on admission. From baseline to discharge, all children received oral micronutrient supplements (vitamins and minerals) that meet WHO specifications for the treatment of children with SAM. Dietary treatment was based on the use of a commercial infant formula-fed (Guigoz 2), containing 498 kcal/100g of energy, 9.9 g/100g of protein, and 23.5 g/100g of fat. The initial feeding phase started with an energy of 75 Kcal per kg per day and 1.2 g protein per kg per day without stopping breastfeeding. Nasogastric tube feeding was used when children did not want to eat or did not have enough appetite or if the child was seriously ill.
The nutritional rehabilitation phase was initiated in malnourished non-edematous children when weight gain was perceived, the infection attenuated and appetite regained. For edematous malnutrition, this nutritional phase was defined clinically by a return of appetite with a substantial loss of edema. The weight loss was accepted as a sign of a positive response to treatment for edematous malnutrition. The frequency and quantity of milk were calculated based on the weight of the child as recommended by the WHO guidelines 9 . Iron supplementation was started when the infection was subsided. In addition, to achieve a consistent intake and rapid weight gain, a highly-sustained diet was maintained during the nutritional rehabilitation phase. During this period, the children were fed a high energy density formula that provided 100-135 kcal per kg per day.

Blood sample
Blood samples were considered part of standard health care. Two blood samples (5 mL) were collected from the radial artery. The first was collected on admission to hospital, within the first 24 hours of hospitalization and before the start of nutritional rehabilitation. While, the second was collected at discharge, after 21 days. The collected blood was centrifuged at 1500 rpm for 15 minutes to separate the sera. The 360 μL quantity of sera was placed in an Eppendorf tube and stored at -20 °C until subsequent nutritional protein analysis. The Hemoglobin concentration was measured by the cyanmethemoglobin method 16 at the biology laboratory of Ibn Khatib Hospital. The determination of the concentrations of acute-phase protein (Acid alpha glycoprotein (AAG) and Creactivate protein (CRP)) was carried out by immunoturbidimetry methods (Hitachi 911, France). The concentration of retinol-binding protein (RBP), transthyretin (TTR), albumin (ALB), transferrin (TRF)) and fibronectin (FN)) were determined by immunoprecipitation in liquid medium using enzyme immunoassay kit on a Dade Behring Bn-100 System Nephelometer. Inflammatory and nutritional proteins were analyzed at the biochemical laboratory, biology department, Faculty of Sciences Dhar Elmahraz Fez.

Anthropometric measurements and Nutritional status assessment
Anthropometric measurements were performed according to the standard technique of the World Health Organization (1976) 17 . At follow-up, children were weighed with minimal clothing in the morning on a scale (RGZ-20), which is accurate to 10 grams. The scale was calibrated every morning before being utilized. Height was measured at admission and discharge by an interchangeable stadiometer (Holtain, Crymych, UK) for standing and recumbent length measurements with the assistance of the dietician. For children < 2 years of age, recumbent length was measured.
Mid-upper-arm circumference (MUAC) was measured with a plastic flexible tape at hospital admission and discharge (21 days), and the readings were recorded at 0.1 cm near. Age was obtained from interviews with mothers. The collected anthropometric measures were converted into weight-forheight z-score (WFH-Z), weight-for-age z-score (WFA-Z), height-for-age z-score (HFA-Z), mid-upper-arm circumference-for-age Z-score (MUAC-Z), and head circumference-for-age Z-score (HC-Z) indices by age and sex using WHO Anthro V3.2.2 for PC 18 . We assessed the nutritional status of the children referring to the WHO criteria for SAM 10 as follows: MUAC less than 11.5cm and/or a WFH-Z less than -3SD or the presence of bipedal pitting edema 10

Nutritional recovery assessment and statistical analysis
The biochemical assessment concerned serum levels of RBP, TTR, ALB, and TRF which have been clinically recognized as the most sensitive proteins used in the assessment of overall nutritional status 20,21 . However, the nutritional and biochemical recovery scheme was assessed in both the edematous and non-edematous groups by calculating the rates of change in serum levels of main nutritional biomarkers (RBP, TTR, ALB, and TRF) between admission and discharge (21-day). Rates of change of major serum nutritional proteins from admission to discharge were calculated as follows: The normality of the data distribution was analyzed by the Kolmogorov-Smirnov test. Rate change data of RBP, TTR, ALB, and TRF, which were not normally distributed, were expressed as the median with a 95% confidence interval (Table 3). However, differences in the rate change of nutritional protein were analyzed by the non-parametric Mann-Whitney U-test, due to their skewed distribution (Rate of nutritional recovery data on table 3). Differences were analyzed with Student's t-test for the normally distributed variables (mean of anthropometric measurements and Z-score indices shown in Tables 1 and 4 and mean of biochemical markers in Table 2). The Chi-Square test was used to compare the categorical variable between the two groups (Difference between age groups and between sex groups in Table 1). The difference was significant when the p-value was < 0.05. All data was analyzed by MedCal (version 9.3.0.0).

Regulatory and ethical aspects
Following the national recommendations of the Ministry of Health, any child who visits a health facility or a hospital is assigned to the assessment of his nutritional status. And any child considered undernourished must be hospitalized to treat their nutritional issue.
This study was conducted in accordance with the Declaration of Helsinki and was approved by the Ethics and Research Committee of the Regional Directorate of the Ministry of Health. The administration of the Ibn Khatib hospital gave its approval to carry out this study. Parents of children who agreed to have their children participate in the study were informed of the purpose and nature of the study. After giving their oral consent for their child's participation, parents were asked to sign a written consent. Mothers were asked to stay with their children during the short hospital stay for nutritional rehabilitation in order to take care of their children and to continue breastfeeding especially at night. If the mothers refused their children's participation in the study, their children had to be hospitalized and receive medical treatment with nutritional rehabilitation as part of the standard treatment.

Results
A total of 25 (25. Table 2 shows the results of nutritional recovery at hospital admission and discharge in edematous and non-edematous children. In both groups, all nutritional proteins and hemoglobin were under the normal range and showed a significant increase to reach their baseline range in the nonedematous and edematous groups except for FN. In contrast, at admission, serum CRP and AAG levels were above normal in both groups and decreased significantly to normal levels at discharge. At discharge, the edematous group showed significantly higher levels of RBP, ALB, TRF and Hg levels than the non-edematous children. As we observed, the rate of recovery of RBP, TTR, and ALB was positive and significantly higher in the edematous than the non-edematous group (p < 0.001), except for TRF which showed an insignificant improvement (p = 0.324).  None: Non-edematous group; Edem: Edematous group; P1: comparison between admission and at discharge in the non-edematous group; P2: comparison between admission and at discharge in the edematous group; P3: comparison between the edematous and non-edematous group upon hospital admission; P4: comparison between the edematous and nonedematous group at discharge; Serum concentration of protein expressed as mean ± standard deviation; The Student T-test considered significant at P-value <0.05.

Discussion
After a short hospital stay for nutritional rehabilitation, we found differences in the pattern of nutritional recovery between children with edematous and non-edematous malnutrition. As shown in Table 3, the magnitude of the response to renutrition was greater in the edematous children than in the non-edematous group, as indicated by the significant increase in recovery rate of nutritional protein (RBP: 115.65 % vs. 44.00; TTR: 154.00 % vs. 89.00; ALB: 98.00 % vs. 10.30; p < 0.001). However, the response to the renutrition was assigned to a different degree between the two groups; the edematous children recovered faster than the nonedematous. These findings are in accordance with previous studies [22][23][24] . Furthermore, previous studies have reported a significant difference in recovery kinetics between edematous and non-edematous malnutrition 25,26 .
More importantly, in the present study, we revealed that at the time of admission, edematous children had significantly lower concentration levels of RBP, TTR, ALB, TRF, and Hg, than the non-edematous group, which has also been demonstrated in previous studies 27,28 . Furthermore, a recent study has revealed serious metabolic disturbance in the edematous child than those without edema 29 . In our study, at discharge, all nutrition-related proteins increased significantly to normal in both groups (Table 4) reflecting satisfactory nutritional recovery except for Hg which remained below normal in the non-edematous group. Additionally, the present study found that after SHS for nutritional rehabilitation, both edematous and nonedematous groups had similar concentrations of RBP, TTR, and FN as reported previously 26 . The repletion of the serum pool of RBP, TTR, ALB, and TRF within the normal range was due to a rapid increase in the rate of synthesis of these proteins in response to adequate protein intake 30,31 .
Moreover, acute-phase protein (APP) study demonstrated that both edematous and non-edematous children had significantly higher serum concentrations of CRP and AAG. These results are in agreement with the study by Reid et al. 32 who showed that both edematous and non-edematous malnourished children had higher plasma concentrations of APP. On the other hand, at the time of admission, there was no significant difference in the CRP and AAG concentrations (40.13 ± 38.67 v.s 55.23 ± 34.04 mg/L for CRP and 8.79 ± 2.54 v.s 29.78 ± 1.66 g/L for AAG) between the edematous and non-edematous, respectively. This result followed that of previous research 33,34 . However, the high CRP and AAG values at admission, in both groups, provide evidence of a response to the infectious process and indicate complicated SAM in these children. After a short hospital stay for nutritional rehabilitation, acute phase proteins decrease significantly to normal in both edematous and nonedematous patients (4.13 ± 1.34 vs. 3.28 ± 1.6 mg/L for CRP and 1.55 ± 0.26 vs. 1.03 ± 0.89 g/L for AAG, respectively). This reduction revealed an attenuation of the inflammatory state and thus testified to the successful clinical recovery. In addition, the return of the serum nutritional protein to normal in both groups provides evidence of a positive response to SHS for nutrition rehabilitation. However, overall, the nutritional recovery profile showed the expected improvement in edematous and non-edematous malnutrition children after SHS. This result is in agreement with recent studies that reported a complete nutritional recovery after a short period of renutrition ranging from 14 to 26 days [35][36][37] .
Furthermore, based on serum protein concentration ( Table  2), we conclude that these children were fully recovered and must return to their home environment after SHS. Yet, the nutritional rehabilitation of malnourished children must allow them to recover a normal nutritional state and to prevent relapses post-discharge in the medium and long term. It should be noted that the anthropometric indicators (Table 4) were still under the WHO recommended discharge criteria 10 for the non-edematous children compared to the edematous group that reached this threshold. The WHO discharge criteria recommends that after nutritional rehabilitation -performed for at least two weeks -malnourished children should reach the threshold of (WFH-Z > -2SD) of the WHO reference population or (MUAC ≥ 12.5 cm) with the absence of nutritional edema 10 .
In the present study, after the SHS, the non-edematous children still had wasting (WFH-Z = -2.02 ± 2.43) than the edematous group (WFH-Z = -1.30 ± 1.05) who met the threshold (WFH-Z > -2SD) ( Table 4). This result is in agreement with the finding of Saaka et al. 24 who showed that non-edematous children stay longer before recovery than their edema counterparts. Furthermore, despite this improvement in WFH-Z, both groups showed persistence of severe stunting (-3.96 ± 2.76 and -3.45 ± 1.78 in the nonedematous and edematous group, successively) ( Table 4). This reflects the insufficient anthropometric recovery after a 21-day of SHS. It is recognized that full catch-up growth is not achieved after a short-term nutritional rehabilitation at the hospital 38 . A study from Jamaica reported that full muscle recovery is slow or impossible after clinical nutritional recovery from SAM 39 . However, in the present study, the biochemical nutritional recovery precedes anthropometric recovery in non-edematous malnutrition children. This result is in agreement with the previous study 37 which emphasizes that the weight/height relationship as a recovery indicator does not coincide with the recovery of other physiological or anthropometric parameters. In light of this, we suggest that early discharge based on biochemical nutritional recovery is possible for both the edematous and non-edematous groups because the children were clinically cured, but not convincing for the non-edematous children because the WHO anthropometric discharge criteria (WFH-  10 were not reached in this group. Given our results, the difference in nutritional recovery between the edematous and non-edematous groups is indicative of intrinsic pathophysiological differences between the two groups (Table 3). We have shown that the response to renutrition differs according to the pathophysiology of SAM as well as within the same group (Table 3). This indicates variability in metabolic and adaptive capacity for each group of children depending on the clinical form of SAM.
It therefore seems to us that the therapeutic approach to the treatment of SAM, proposed by the WHO, should take into consideration the physiological differences between the edematous and non-edematous forms. However, further research is needed to understand the nutritional recovery pattern and physiological abnormalities in edematous and non-edematous malnutrition and their response to renutrition. These detailed clinical studies performed in these two forms of malnutrition with precisions on metabolic status, clinical status and response to different nutritional therapies, during the first period of nutritional rehabilitation in the hospital, will be an essential basis for offering specific management and clinical practices for each type of SAM.
Like any research study, some limitations were unavoidable.
In the literature review, we encountered the heterogeneity of the protocols used to treat malnourished children. All previous studies were carried out using different types of dietary treatment (protein, energy, and micronutrients), duration of treatment, admission and discharge criteria, definitions used for SAM, and age of children studied. This heterogeneity made it difficult to compare the results of our study. In addition, the study's small sample size is an important limitation. The small sample size limits the generalization of our results and is due to the reduced prevalence of SAM in our country. However, further studies with a large sample size in other different populations are needed. Furthermore, in our study, we did not investigate relapse after discharge. Proposed WHO guidelines for the treatment of children with severe acute malnutrition recommend follow-up of children after discharge from hospital to prevent relapses.

Conclusions
Our study found that the pattern of nutritional recovery appeared to differ between edematous and non-edematous malnutrition. The edematous child seems to recover faster than their non-edematous counterparts. Furthermore, normalization of the serum pool of RBP, TTR, ALB, TRF, and attenuation of inflammatory status indicated by decreases in CRP and AAG, after a short-term inpatient nutritional rehabilitation program of 21 days, provides evidence of complete biochemical nutritional recovery in edematous and non-edematous malnutrition. In addition, at discharge, despite complete biochemical nutritional recovery, severe stunting persisted in both groups, with the presence of moderate wasting in the non-edematous group compared with the edematous group, which met WHO anthropometric criteria for discharge from nutritional programs.
Source(s) of support: This research was supported by the Ministry of Higher Education and Innovation, Morocco, project PROTARS P1T1/05. Acknowledgment: We are particularly indebted to pediatric staff and we thank to dietician and nurses for their personal involvement in this study. We also acknowledge the director and staff of laboratory of biological analysis at Ibn Khatib Hospital, for their co-operation and assistance.