Changes in polyunsaturated fatty acids during treatment of malnourished children may be insufficient to reach required essential fatty acid levels - A randomised controlled trial.

BACKGROUND & AIMS: Severe acute malnutrition (SAM) is a global concern. Studies on the impact of ready-to-use therapeutic foods (RUTFs) on polyunsaturated fatty acids (PUFA) are almost non-existent. The aim was to investigate the change in whole-blood PUFA and nutrition and health markers among Cambodian children with SAM after treatment with RUTFs. METHODS: The trial was an 8-week randomised clinical trial of the effectiveness of locally produced fish-based RUTF (L-RUTF) vs standard milk-based RUFT (S-RUTF). Whole-blood fatty acids were analysed using dried blood spots. Nutrition and health markers were assessed using anthropometric assessment and blood samples for markers of inflammation. The trial was conducted at the National Pediatric Hospital, Phnom Penh, Cambodia, with one hundred and twenty-one 6-59-month-old children in treatment for SAM. RESULTS: L-RUTF had a higher content of n-3 PUFA and a higher content of arachidonic acid (AA) and docosahexaenoic acid (DHA), while S-RUTF had the highest content of n-6 PUFA. At baseline, the children presented with a Mead acid level in whole-blood of around 0.08% of total fatty acids (FA%) and an omega-3 index of ∼0.91 ± 0.44. After eight weeks of S-RUTF treatment, linoleic acid (LA), AA, n-6/n-3 PUFA ratio, and Mead acid levels were increased. The L-RUTF intervention did not change the whole-blood PUFAs from baseline. At discharge, the children in the L-RUTF group had a lower n-6/n-3 PUFA ratio than the children in the S-RUTF group, driven by a lower alpha-linolenic acid (ALA) (0.20 vs 0.27 FA%, p = 0.004) and lower LA (15.77 vs 14.21 FA%, p = 0.018) with no significant differences in AA or DHA levels. Weight-for-height z-score at discharge was negatively associated with total PUFA (ß -1.4 FA%, 95%CI. -2.7; -0.1), n-6 LCPUFA (ß -1.3 FA%, 95%CI. -1.3; -0.3), and AA (ß -0.6 FA%, 95%CI. -1.0; -0.2). Age-adjusted height was negatively associated with the Mead acid:AA ratio (ß -1.2 FA%, 95%CI. -2.2; -0.2). No significant change was seen in inflammation markers within groups or between groups during treatment, and n-3 and n-6 PUFAs were not associated with markers of inflammation or haemoglobin status at discharge. CONCLUSION: The trial found that whole-blood markers of PUFA status were low in children at admission and discharge from SAM treatment, indicating that the currently recommended composition of RUTFs are not able to correct their compromised essential fatty acid status. The higher content of DHA and AA in L-RUTF did not give rise to any improvement in PUFA status. No changes in health markers or associations between PUFA and health markers were found. TRIAL REGISTRATION: ClinicalTrials.gov: NCT02907424.

Whole-blood PUFA and associations with markers of nutritional and health status in acutely malnourished children in Cambodia.

OBJECTIVE: To measure fatty acid composition, particularly whole-blood PUFA content, in acutely malnourished children and identify associations with markers of nutritional and health status. DESIGN: PUFA were assessed in dried blood spots obtained from a cross-sectional study. Nutritional and health status were assessed by anthropometry, haemoglobinopathies, inflammation and blood counts. SETTING: Cambodia. PARTICIPANTS: The study was conducted with 174 children aged 0·5-18 years with acute malnutrition. RESULTS: Among total fatty acids (FA), the relative percentage of total PUFA was 20 % FA, with 14 % of the children having very low PUFA (mead acid (MA):arachidonic acid (AA) >0·02, n-6 docosapentaenoic acid:DHA >0·2 and total n-6:n-3 PUFA >10·5). Wasting was not associated with any PUFA. Stunting and low height were consistently positively associated with total PUFA and positively with n-6 PUFA. Height was positively associated with n-3 long-chain PUFA (LCPUFA). The presence of haemoglobinopathies or inflammation was positively associated with MA:AA, but not total PUFA. Elevated blood platelet counts were positively correlated with linoleic acid and appeared to be influenced by anaemia (P = 0·010) and inflammation (P = 0·002). Monocyte counts were high during inflammation (P = 0·052) and correlated positively with n-6 LCPUFA and n-3 LCPUFA. CONCLUSIONS: Children with acute malnutrition or stunting had low PUFA, while elevated platelets and monocytes were associated with high PUFA. In acutely malnourished children, inflammation could lead to elevated blood cell counts resulting in increased whole-blood PUFA which does not reflect dietary intake or nutritional status.

Effectiveness of a locally produced ready-to-use supplementary food in preventing growth faltering for children under 2 years in Cambodia: a cluster randomised controlled trial.

This cluster randomised controlled trial tested the effectiveness of a locally produced, fish-based, ready-to-use supplementary food (RUSF) to prevent growth faltering (decline in z-scores). Cambodian infants (n= 485), aged 6 to 11 months, were randomised by site to receive the RUSF, Corn-Soy Blend++ (CSB++), micronutrient powders (MNP), or no supplement (control). The intervention was for 6 months. In unadjusted analysis, the control group had statistically significantly decreased weight-for-age z-scores (WAZ; -0.02, 95%CI = -0.03 - -0.01, P= 0.001) and height-for-age z-scores (HAZ; -0.07, 95%CI = -0.09 - -0.05, P < 0.001), and increased mid-upper arm-circumference (MUAC; 0.02cm, 95%CI = 0.01 - 0.04, P = 0.010), but no statistically significant change in weight-for-height z-scores (WHZ). The RUSF group did not differ significantly from the control for WAZ, HAZ or WHZ (in other words, WAZ and HAZ decreased and WHZ did not change), but had increased MUAC in comparison to the control (0.04cm, 95%CI = 0.01 - 0.06, P = 0.008). There were no statistically significant differences between the RUSF group and the CSB++ or MNP groups with respect to WAZ, HAZ, WHZ or MUAC. Interestingly, in adjusted analysis, low consumers of RUSF had increased WAZ, WHZ and MUAC (0.03, 95%CI = 0.01-0.06, P = 0.006; 0.04, 95%CI = 0.01-0.08, P = 0.026; and 0.05cm, 95%CI = 0.02-0.09, P = 0.004, respectively) compared with the control. The novel RUSF, particularly in small quantities, protected against ponderal growth faltering, but the improvements were of limited clinical significance.

Multi-Micronutrient Fortified Rice Improved Serum Zinc and Folate Concentrations of Cambodian School Children. A Double-Blinded Cluster-Randomized Controlled Trial.

BACKGROUND: Within Cambodia, micronutrient deficiencies continue to be prevalent in vulnerable groups, such as women and children. Fortification of staple foods such as rice could be a promising strategy for Cambodia to improve micronutrient status. OBJECTIVE: Our objective was to investigate the impact of multiple-micronutrient fortified rice (MMFR), distributed through a World Food Program school-meals program (WFP-SMP) on serum zinc concentrations and folate status in a double-blind, cluster-randomized, placebo-controlled trial. METHODS: Sixteen schools were randomly assigned to receive one of three different types of extruded-fortified rice (UltraRice Original (URO), UltraRice New (URN), or NutriRice) or unfortified rice (placebo) six days a week for six months. A total of 1950 schoolchildren (6-16 years old) participated in the study. Serum zinc (all groups) and folate (only in NutriRice and placebo group) concentrations were assessed from morning non-fasting antecubital blood samples and were measured at three time points (baseline and after three and six months). RESULTS: After six months of intervention, serum zinc concentrations were significantly increased in all fortified rice group compared to placebo and baseline (0.98, 0.85 and 1.40 µmol/L for URO, URN and NutriRice, respectively) (interaction effect: p < 0.001 for all). Children in the intervention groups had a risk of zinc deficiencies of around one third (0.35, 039, and 0.28 for URO, URN, and NutriRice, respectively) compared to the placebo (p < 0.001 for all). The children receiving NutriRice had higher serum folate concentrations at endline compared to children receiving normal rice (+ 2.25 ng/mL, p = 0.007). CONCLUSIONS: This study showed that the high prevalence of zinc and folate deficiency in Cambodia can be improved through the provision of MMFR. As rice is the staple diet for Cambodia, MMFR should be considered to be included in the school meal program and possibilities should be explored to introduce MMFR to the general population.

Development and testing of locally-produced ready-to-use therapeutic and supplementary foods (RUTFs and RUSFs) in Cambodia: lessons learned.

BACKGROUND: Rates of childhood undernutrition are persistently high in Cambodia. Existing ready-to-use supplementary and therapeutic foods (RUSFs and RUTFs) have had limited acceptance and effectiveness. Therefore, our project developed and trialled a locally-produced, multiple micronutrient fortified lipid-based nutrient supplement (LNS) with therapeutic and supplementary versions. This ready-to-use food (RUF) is innovative in that, unlike many RUFs, it contains fish instead of milk. Development began in 2013 and the RUF was finalised in 2015. From 2015 until the present, both the RUTF and the RUSF versions were trialled for acceptability and effectiveness. METHODS: This paper draws on project implementation records and semi-structured interviews to describe the partnership between the Cambodian Ministries of Health and Agriculture, Forestry and Fisheries, UNICEF, the French National Research Institute for Sustainable Development (IRD), universities, and Vissot factory. It discusses the project implementation and lessons learned from the development and trialling process, and insights into positioning nutrition on the health agenda in low and middle-income countries. RESULTS: The lessons learned relate to the importance of project planning, management, and documentation in order to seize opportunities in the research, policy, advocacy, and programming environment while ensuring adequate day-to-day project administration and resourcing. CONCLUSIONS: We conclude that projects such as ours, that collaborate to develop and test novel, locally-produced RUTFs and RUSFs, offer an exciting opportunity to respond to both local programmatic and broader research needs.

Multi-criteria Mapping of Stakeholders' Viewpoints in Five Southeast Asian Countries on Strategies to Reduce Micronutrient Deficiencies Among Children and Women of Reproductive Age: Findings from the SMILING Project.

BACKGROUND: Despite scientific evidence on the potential impact or importance of specific interventions to improve micronutrient status of vulnerable groups, political commitment and extensive support from national stakeholders is paramount to support introduction and implementation of these interventions at national level. In order to develop efficient nutritional strategies to improve the micronutrient status of children < 5 years of age and women of reproductive age that will be supported by a wide range of stakeholders, a better understanding of viewpoints on the nutrition politics and strategies is necessary. Multi-criteria mapping (MCM) was successfully used to assess the stakeholder's viewpoint in a wide variety of contexts since the late 1990s. OBJECTIVE: The objective of the present study was to assess the viewpoints of stakeholders on a wide range of potential nutritional interventions in the five Southeast Asian countries participating in the SMILING project. METHOD: MCM methodology was used to appraise the stakeholder's viewpoints in five countries. RESULTS: The results show that the overall stakeholders' preference was for actions already implemented in their country rather than for new, innovative options, even for supplementation. Indirect interventions such like food fortification (except in Indonesia), delayed cord clamping or food-based approaches were generally less favored by the stakeholders. However, the majority of stakeholders agreed that new approaches should be considered and put in place in the future provided that evidence of their impact was demonstrated, that they received adequate technical support for their implementation and their monitoring, and that they will be accompanied by strong advocacy among decision-makers, civil society and beneficiaries. CONCLUSIONS FOR PRACTICE: To conclude, for the introduction of new, innovative strategies to reduce micronutrient deficiencies in South-East Asia, convincing stakeholders appears to be the first hurdle to be taken.

Effect of multi-micronutrient-fortified rice on cognitive performance depends on premix composition and cognitive function tested: results of an effectiveness study in Cambodian schoolchildren.

OBJECTIVE: Even though current policy is strongly focused on the crucial first '1000 days', it might be still possible to enhance cognitive function during the pre-adolescent and adolescent years by improving micronutrient status. In Cambodia, nutritional status is poor. Provision of rice fortified with micronutrients through a school meal programme (SMP) could be a cost-effective strategy to help improve health and school performance. The present study aimed to evaluate the effect of three different micronutrient-fortified rice formulations on cognitive function in Cambodian children. SETTING: Sixteen Cambodian schools receiving SMP. DESIGN: The FORISCA-UltraRice®+NutriRice® study was a randomized, double-blind, placebo-controlled trial. Four groups of four schools were randomly allocated to receive normal rice, UltraRice®Original, UltraRice®New or NutriRice®. Within each school, 132 children were randomly selected. Data on cognitive performance (picture completion, block design and Raven's coloured progressive matrices (RCPM)), anthropometry, parasite infestation and micronutrient status were collected before the intervention and after 6 months. SUBJECTS: Cognitive data were available for 1796 children aged 6-16 years. RESULTS: All cognitive scores improved after 6 months (P<0·001). Block design score improvement was significantly higher in children consuming UltraRice®Original (P=0·03) compared with the other fortified rice groups and placebo. No difference among groups was found on RCPM or picture completion scores. Stunting, parasite infestation and inflammation negatively affected the impact of the intervention. CONCLUSIONS: Combined with other interventions, using SMP to distribute fortified rice to schoolchildren may be a cost-effective way to increase cognitive performance and thereby improve school performance and educational achievements.

Crossover trial to test the acceptability of a locally produced lipid-based nutrient supplement (LNS) for children under 2 years in Cambodia: a study protocol.

INTRODUCTION: The acceptability and efficacy of existing ready-to-use supplementary and therapeutic foods has been low in Cambodia, thus limiting success in preventing and treating malnutrition among Cambodian children. In that context, UNICEF and IRD have developed a locally produced, multiple micronutrient fortified lipid-based nutrient supplement. This food is innovative, in that it uses fish instead of milk as the animal source food. Very few supplementary foods have non-milk animal source foods, and in addition they have not been widely tested. This trial will assess the novel food's acceptability to children and caregivers. METHODS AND ANALYSIS: This is a cluster-randomised, incomplete block, 4×4 crossover design with no blinding. It will take place in four sites in a community setting in periurban Phnom Penh. Healthy children aged 9-23 months (n=100) will eat each of four foods for 3 days at a time. The amount they consume will be measured, and at the end of each 3-day set, caregivers will assess how well their child liked the food. After 12 days, caregivers themselves will do a sensory test of the 4 foods and will rank them in terms of preference. ETHICS AND DISSEMINATION: Ethical clearance was received from the University of Queensland Medical Research Ethics Committee (2014001070) and from Cambodia's National Ethics Committee for Health Research (03/8 NECHR). REGISTRATION: ClinicalTrials.gov, identifier: LNS-CAMB-INFANTS; NCT02257437. Pre-results.

Low Urinary Iodine Concentration among Mothers and Children in Cambodia.

A 2014 national assessment of salt iodization coverage in Cambodia found that 62% of samples were non-iodized, suggesting a significant decline in daily iodine intakes. The Cambodian Micronutrient Survey conducted in 2014 (CMNS-2014) permitted obtaining national data on urinary iodine concentrations (UIC) to assess iodine status and whether iodized salt use had an impact. Urine samples were collected from mothers (n = 736) and children (n = 950). The median UIC was 63 µg/L and 72 µg/L in mothers and children respectively. More than 60% of mothers and their children had a UIC < 100 µg/L, thereby indicating a serious public health problem. Iodine status was significantly lower among mothers and children living in rural areas, belonging to the poorest socioeconomic category, or living in a household not using iodized salt. The limited enforcement of the legislation for iodized salt has resulted in a major decrease in the prevalence of iodized salt, which in turn has compromised iodine status in Cambodia. It is essential for the government to enhance enforcement of the iodized salt legislation, and implement short term strategies, such as iodine supplementation, to prevent an increase of severe complications due to iodine deficiency in the Cambodian population.

Impact of Multi-Micronutrient Fortified Rice on Hemoglobin, Iron and Vitamin A Status of Cambodian Schoolchildren: a Double-Blind Cluster-Randomized Controlled Trial.

In Cambodia, micronutrient deficiencies remain a critical public health problem. Our objective was to evaluate the impact of multi-micronutrient fortified rice (MMFR) formulations, distributed through a World Food Program school-meals program (WFP-SMP), on the hemoglobin concentrations and iron and vitamin A (VA) status of Cambodian schoolchildren. The FORISCA-UltraRice+NutriRice study was a double-blind, cluster-randomized, placebo-controlled trial. Sixteen schools participating in WFP-SMP were randomly assigned to receive extrusion-fortified rice (UltraRice Original, UltraRice New (URN), or NutriRice) or unfortified rice (placebo) six days a week for six months. Four additional schools not participating in WFP-SMP were randomly selected as controls. A total of 2440 schoolchildren (6-16 years old) participated in the biochemical study. Hemoglobin, iron status, estimated using inflammation-adjusted ferritin and transferrin receptors concentrations, and VA status, assessed using inflammation-adjusted retinol-binding protein concentration, were measured at the baseline, as well as at three and six months. Baseline prevalence of anemia, depleted iron stores, tissue iron deficiency, marginal VA status and VA deficiency were 15.6%, 1.4%, 51.0%, 7.9%, and 0.7%, respectively. The strongest risk factors for anemia were hemoglobinopathy, VA deficiency, and depleted iron stores (all p < 0.01). After six months, children receiving NutriRice and URN had 4 and 5 times less risk of low VA status, respectively, in comparison to the placebo group. Hemoglobin significantly increased (+0.8 g/L) after three months for the URN group in comparison to the placebo group; however, this difference was no longer significant after six months, except for children without inflammation. MMFR containing VA effectively improved the VA status of schoolchildren. The impact on hemoglobin and iron status was limited, partly by sub-clinical inflammation. MMFR combined with non-nutritional approaches addressing anemia and inflammation should be further investigated.

Stability of Vitamin A, Iron and Zinc in Fortified Rice during Storage and Its Impact on Future National Standards and Programs--Case Study in Cambodia.

Fortified rice holds great potential for bringing essential micronutrients to a large part of the world population. The present study quantified the losses of three different micronutrients (vitamin A, iron, zinc) in fortified rice that were produced using three different techniques (hot extrusion, cold extrusion, and coating) and stored at two different environments (25 ± 5 °C at a humidity of 60% and 40 ± 5 °C at a humidity of 75%) for up to one year. Fortified rice premix from the different techniques was mixed with normal rice in a 1:100 ratio. Each sample was analyzed in triplicate. The study confirmed the high stability of iron and zinc during storage while the retention of vitamin A was significantly affected by storage and the type of techniques used to make rice premix. Losses for iron and zinc were typically <10% for any type of rice premix. After 12 months at mild conditions (25 °C and humidity of 60%), losses for vitamin A ranged from 20% for cold extrusion, 30% for hot extruded rice 77% for coated rice premix. At higher temperatures and humidity, losses of vitamin A were 40%-50% for extruded premix and 93% for coated premix after 6 months. We conclude that storage does lead to a major loss of vitamin A and question whether rice is a suitable food vehicle to fortify with vitamin A. For Cambodia, fortification of rice with iron and zinc could be an effective strategy to improve the micronutrient status of the population if no other food vehicles are available.

Micronutrient-Fortified Rice Can Increase Hookworm Infection Risk: A Cluster Randomized Trial.

BACKGROUND: Fortification of staple foods is considered an effective and safe strategy to combat micronutrient deficiencies, thereby improving health. While improving micronutrient status might be expected to have positive effects on immunity, some studies have reported increases in infections or inflammation after iron supplementation. OBJECTIVE: To study effects of micronutrient-fortified rice on hookworm infection in Cambodian schoolchildren. METHODS: A double-blinded, cluster-randomized trial was conducted in 16 Cambodian primary schools partaking in the World Food Program school meal program. Three types of multi-micronutrient fortified rice were tested against placebo rice within the school meal program: UltraRice_original, UltraRice_improved and NutriRice. Four schools were randomly assigned to each study group (placebo n = 492, UltraRice_original n = 479, UltraRice_improved n = 500, NutriRice n = 506). Intestinal parasite infection was measured in fecal samples by Kato-Katz method at baseline and after three and seven months. In a subgroup (N = 330), fecal calprotectin was measured by ELISA as a marker for intestinal inflammation. RESULTS: Baseline prevalence of hookworm infection was 18.6%, but differed considerably among schools (range 0%- 48.1%).Micronutrient-fortified rice significantly increased risk of new hookworm infection. This effect was modified by baseline hookworm prevalence at the school; hookworm infection risk was increased by all three types of fortified rice in schools where baseline prevalence was high (>15%), and only by UltraRice_original in schools with low baseline prevalence. Neither hookworm infection nor fortified rice was related to fecal calprotectin. CONCLUSIONS: Consumption of rice fortified with micronutrients can increase hookworm prevalence, especially in environments with high infection pressure. When considering fortification of staple foods, a careful risk-benefit analysis is warranted, taking into account severity of micronutrient deficiencies and local prevalence of parasitic infections. TRIAL REGISTRATION: ClinicalTrials.gov NCT01706419.

The use of linear programming to determine whether a formulated complementary food product can ensure adequate nutrients for 6- to 11-month-old Cambodian infants.

BACKGROUND: A new software tool, Optifood, developed by the WHO and based on linear programming (LP) analysis, has been developed to formulate food-based recommendations. OBJECTIVE: This study discusses the use of Optifood for predicting whether formulated complementary food (CF) products can ensure dietary adequacy for target populations in Cambodia. DESIGN: Dietary data were collected by 24-h recall in a cross-sectional survey of 6- to 11-mo-old infants (n = 78). LP model parameters were derived from these data, including a list of foods, median serving sizes, and dietary patterns. Five series of LP analyses were carried out to model the target population's baseline diet and 4 formulated CF products [WinFood (WF), WinFood-Lite (WF-L), Corn-Soy-Blend Plus (CSB+), and Corn-Soy-Blend Plus Plus (CSB++)], which were added to the diet in portions of 33 g/d dry weight (DW) for infants aged 6-8 mo and 40 g/d DW for infants aged 9-11 mo. In each series of analyses, the nutritionally optimal diet and theoretical range, in diet nutrient contents, were determined. RESULTS: The LP analysis showed that baseline diets could not achieve the Recommended Nutrient Intake (RNI) for thiamin, riboflavin, niacin, folate, vitamin B-12, calcium, iron, and zinc (range: 14-91% of RNI in the optimal diets) and that none of the formulated CF products could cover the nutrient gaps for thiamin, niacin, iron, and folate (range: 22-86% of the RNI). Iron was the key limiting nutrient, for all modeled diets, achieving a maximum of only 48% of the RNI when CSB++ was included in the diet. Only WF and WF-L filled the nutrient gap for calcium. WF-L, CSB+, and CSB++ filled the nutrient gap for zinc (9- to 11-mo-olds). CONCLUSIONS: The formulated CF products improved the nutrient adequacy of complementary feeding diets but could not entirely cover the nutrient gaps. These results emphasize the value of using LP to evaluate special CF products during the intervention planning phase. The WF study was registered at controlled-trials.com as ISRCTN19918531.

Isolation and identification of 3-methylcrotonyl coenzyme A carboxylase cDNAs and pyruvate carboxylase, and their expression in red seabream (Pagrus major) organs.

We determined complementary DNA sequences of biotin-containing (MCCC1) and non-biotin-containing (MCCC2) subunits of 3-methylcrotonyl coenzyme A carboxylase (MCCase) and pyruvate carboxylase (PCase) using reverse transcriptase polymerase chain reaction of RNA extracted from seabream skeletal muscle and liver. We determined the complete coding sequences of MCCC1 and PC and a partial coding sequence of the major part of MCCC2. Molecular sizes of MCCC1, MCCC2, and PC were 4300, 2400, and 6500 nucleotides, respectively, according to Northern blot analysis. The length of MCCC1 from cDNA sequencing was 4249 nucleotides, indicating the full-length messenger RNA sequence was obtained. Northern blot analyses showed that PC was expressed in muscle, heart, liver, and ovary, but not in spleen. MCCC1 and MCCC2 were expressed at high levels in muscle and ovary, but only trace levels in heart, spleen, and liver. MCCase appears to be particularly important in muscle and ovary, which are active in protein metabolism, while PCase is important in organs active in glycolysis, such as liver.