AJCN. First published ahead of print January 28, 2015 as doi: 10.3945/ajcn.114.084889.
Effects of animal source food and micronutrient fortification in
complementary food products on body composition, iron status, and
linear growth: a randomized trial in Cambodia1–4
Jutta KH Skau, Bunthang Touch, Chamnan Chhoun, Mary Chea, Uma S Unni, Jan Makurat, Suzanne Filteau,
Frank T Wieringa, Marjoleine A Dijkhuizen, Christian Ritz, Jonathan C Wells, Jacques Berger, Henrik Friis,
Kim F Michaelsen, and Nanna Roos
ABSTRACT
Background: Poor nutritional quality of complementary foods often limits growth. Animal source foods, such as milk or meat, are
often unaffordable. Local affordable alternatives are needed.
Objective: We evaluate the efficacy of 2 newly developed, ricebased complementary food products: WinFood (WF) with small fish
and edible spiders and WinFood-Lite (WF-L) fortified with small
fish, against 2 existing fortified corn-soy blend products, CSB+
(purely plant based) and CSB++ (8% dried skimmed milk).
Design: In total, 419 infants aged 6 mo were enrolled in this randomized, single-blinded study for 9 mo, designed primarily to assess increments in fat-free mass by a deuterium dilution technique
and change in plasma ferritin and soluble transferrin receptor. Secondary endpoints were changes in anthropometric variables, including knee-heel length. Data were analyzed by the intention-to-treat
approach.
Results: There was no difference in fat-free mass increment in WF
or WF-L compared with CSB+ [WF: +0.04 kg (95% CI: 20.20,
0.28); WF-L: +0.14 kg (95% CI: 20.10, 0.38)] or CSB++ [WF:
20.03 kg (95% CI: 20.27, 0.21); WF-L: +0.07 kg (95% CI: 20.18,
0.31)] and no effect on the iron status. The 1.7 mm (95% CI: 20.1,
3.5) greater increase in knee-heel length in WF-L compared with
CSB+ was not significant.
Conclusions: No difference was found between the locally produced products (WF and WF-L) and the CSBs. Micronutrient fortification may be necessary, and small fish may be an affordable
alternative to milk to improve complementary foods. The dietary
role of edible spiders needs to be further explored. This trial was
registered at controlled-trials.com as ISRCTN19918531.
Am J
Clin Nutr doi: 10.3945/ajcn.114.084889.
Keywords animal source foods, body composition, complementary food, iron status, fortification
INTRODUCTION
The complementary feeding period is well recognized as
a “window of opportunity” for preventing malnutrition (1, 2).
The complementary foods in food-insecure settings typically
suffer from lack of diversity and animal source foods (ASFs),5
resulting in nutritional insufficiency. Lack of evidence from
well-conducted studies, however, limits efficient intervention
strategies (3).
ASFs such as milk and meat are generally beneficial to promote
growth (4) but often unaffordable to food-insecure households.
Small fish are nutritionally beneficial because they are eaten
whole, with head, bones, and viscera (5, 6), and edible insects are
an untapped food resource (7). Food distribution in programs for
the treatment and prevention of malnutrition in infants and
children commonly includes fortified blended food products,
such as corn-soy blends (CSBs) (8). In 2010, the World Food
Programme (WFP) introduced 2 improved formulas of CSB: CSB
plus (CSB+) and CSB plus plus (CSB++). The formula for the
1
From the Department of Nutrition, Exercise and Sports, University of
Copenhagen, Denmark (JKHS, MD, CR, HF, KFM, and NR); Inland Fisheries
Research and Development Institute, Phnom Penh, Cambodia (TB); Department of Fisheries Post-harvest Technologies and Quality Control, Fishery
Administration, Ministry of Agriculture, Forestry and Fisheries, Phnom Penh,
Cambodia (CC); National Nutrition Programme, Maternal and Child Health
Centre, Ministry of Health, Phnom Penh, Cambodia (CM); Institute of Nutritional Sciences, Justus Liebig University, Giessen, Germany (JM); Faculty of
Epidemiology and Population Health, London School of Hygiene and Tropical Medicine, London, United Kingdom (SF); Childhood Nutrition Research
Centre, UCL Institute of Child Health, London, United Kingdom (JW); St.
John’s Research Institute, Bangalore, India (USU); and Institut de Recherche
pour le Développement (IRD), Montpellier, France (FW and JB).
2
Supported by the Consultative Research Committee for Development
Research, Ministry of Foreign Affairs, Copenhagen, Denmark. The Institut
de Recherche pour le Développement donated a grant to the production of
WinFood-Lite. Fortitech Gadstrup donated the calcium phosphate for the
WinFood-Lite. CSB++ was donated by the World Food Programme, Rome, Italy.
CSB+ was donated by World Food Programme, Phnom Penh, Cambodia. The
deuterium dilution technique for measuring body composition was supported with
financial and technical support by International Atomic and Energy Agency.
3
Supplemental Tables 1 and 2 are available from the “Supplemental data”
link in the online posting of the article and from the same link in the online
table of contents at http://ajcn.nutrition.org.
4
Address correspondence to JKH Skau, Department of Nutrition, Exercise
and Sports, University of Copenhagen, Rolighedsvej 30, 1958 Frederiksberg
C, Denmark. E-mail: [email protected].
5
Abbreviations used: ASF, animal source food; CSB, corn-soy blend;
CSB+, corn-soy blend plus; CSB++, corn-soy blend plus plus; FFM, fat-free
mass; FM, fat mass; LAZ, length-for-age z score; MUAC, midupper arm
circumference; sTfR, soluble transferrin receptor; WAZ, weight-for-age
z score; WF, WinFood; WF-L, WinFood-Lite; WFP, World Food Programme;
WLZ, weight-for-length z score.
Received January 29, 2014. Accepted for publication December 23, 2014.
doi: 10.3945/ajcn.114.084889.
Am J Clin Nutr doi: 10.3945/ajcn.114.084889. Printed in USA. Ó 2015 American Society for Nutrition
Copyright (C) 2015 by the American Society for Nutrition
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SKAU ET AL.
mineral and vitamin mix was improved for CSB+ (9), and CSB++
also contains 8% dried skimmed milk and dehulled soya and
soybean oil (10). CSB++ is targeted treatment of moderate malnutrition, and the price is more than double that of CSB+.
This current study was conducted in a food-insecure rural
population in Cambodia, where national data from 2010 showed
that 40% of children aged ,5 y were stunted and 11% were
wasted. Anemia is also highly prevalent in children aged ,5 y in
Cambodia (11). In 2010, the WFP was supporting the food
distribution of CSB+ to food-insecure households and CSB++ to
moderately malnourished children in Cambodia. Locally produced alternatives to the imported CSB products were considered potentially feasible for program implementation. Therefore,
the WinFood products were developed and tested in a randomized trial as possible replacements for CSB+ in more general
programs or for CSB++ in targeted programs. The trial was
a part of the WinFood project, within which nutritionally optimized complementary food products based on local foods were
developed. Culturally and nutritionally feasible WinFood products were developed based on the traditional rice-porridge
borbor, with small fish and edible spiders. Two compositions
were developed, one product fortified with minerals and vitamins, similar to CSB, and one unfortified but with more fish and
edible spiders to contribute micronutrients, especially iron and
zinc.
The study was also designed to assess the effect of complementary feeding beyond the standard anthropometric measures of
height and weight to improve our understanding of the true
impact on healthy growth (12). Body composition was measured,
assessing growth in fat-free mass (FFM) and fat mass (FM). Iron
status was also a primary outcome because improved iron status
in infancy can have long-term effects (13). The objective of the
study was to evaluate these WinFood products as replacement
products for either the standard CSB+ product or the improved
CSB++ product with milk, recognized as the “best product” in
food aid targeting moderate malnutrition. We therefore designed
the study to test whether the impact of WinFood products was
superior to CSB+ and not different from CSB++, because these
selected comparisons would provide answers about the suitability
of WinFood products as a replacement of either CSB product in
complementary feeding in Cambodia.
METHODS
Study participants and setting
The study was conducted from March 2011 to March 2012. All
single-born infants who had turned or would turn age 6 mo in the
following recruitment month from all villages in 7 communes in
PeaRieng and Sithor Kandal Districts, Prey Veng province,
Cambodia, were invited for screening at the referral hospital in
PeaRieng town. Infants were seen by a pediatrician and screened
for severe malnutrition [,23 weight-for-length z score (WLZ)],
pitting edema, clinical signs of vitamin A deficiency, and severe
anemia (hemoglobin ,80 g/L). If any of these symptoms were
detected, the infant was excluded and referred for treatment.
Infants with a history of persistent diarrhea before the time of
recruitment were referred for treatment and invited for a new
screening 2–4 wk later. All caregivers of participating infants
voluntarily signed the consent form for participation and were
informed that they could leave the study at any time. The protocol
was approved by the National Ethics Committee for Health Research, Ministry of Health, the Royal Government of Cambodia
(151 NEHR), and the consultative approval was obtained from the
Danish National Committee on Biomedical Research Ethics. This
trial was registered at controlled-trials.com as ISRCTN19918531.
Study design
This was an individual, randomized, single-blinded, communitybased trial designed to assess change in body composition with
increments in FFM and change in iron status, measured by ferritin
and transferrin receptors (sTfRs), as primary outcomes in infants
receiving one of 4 processed complementary food products: WF
with total 14% dry weight ASF, WF-L with 10% dry weight ASF
and fortified with a mineral and vitamin mix (same as in the 2 CSB
products), CSB+, or CSB++ in a 9-mo period (Table 1 and Table
2). Secondary outcomes were change in length, knee-heel length,
weight, midupper arm circumference (MUAC), head circumference, skinfolds, and length-for-age z score (LAZ), weight-for-age
z score (WAZ), and WLZ. The daily rations of foods before cooking
were 50 g at 6–8 mo of age, 75 g at 9–11 mo of age, and 125 g at
12–15 mo of age. These rations were adjusted to WHO recommendations for complementary feeding of breastfed infants and
supplied ;200, 300, or 550 kcal/d for the 3 age groups, respectively (14). Food distribution and anthropometric measurements took place monthly at health centers in each commune. At
each visit, the date for the next visit was set. The number of days
between monthly follow-up visits varied by 65 d. At each food
distribution, the caregivers were instructed in how to prepare the
food and received an educational session with messages on good
food preparation, food hygiene, good complementary feeding, and
breastfeeding practices, in line with guidelines communicated by
the Ministry of Health, Cambodia.
TABLE 1
Food composition of the intervention foods per 100 g dry weight1
Food composition
Rice, white, milled
Fish, Esomus longimanus4
Fish, Paralaubuca typus4
Spider, Haplopelma species4
Fish mix4
Mineral and vitamin mix
Vegetable oil
Sugar
Maize (white or yellow)
Dehulled soya
Whole soya
Skimmed milk powder
WF,2 % WF-L,2 % CSB++, % CSB+,3 %
77
6.1
6.1
1.8
—
—
4.8
4.8
—
—
—
—
79
—
—
—
9.5
1.7
4.8
4.8
—
—
—
—
—
—
—
—
—
1.7
3.0
9.0
58
20
—
8
—
—
—
—
—
1.4
8.5
8.5
65
—
20
—
1
Food groups: CSB+, corn-soy blend plus; CSB++, corn-soy blend plus
plus; WF, WinFood; WFP, World Food Programme; WF-L, WinFood-Lite.
2
WF and WF-L products were precooked by extrusion, similar to the
processing of CSB products.
3
Included sugar added by the WFP in Cambodia and oil distributed in
separate sachets to be added to the daily rations following WFP product
specifications (version 1.1) (9).
4
Edible parts of the fish were used, obtained by traditional cleaning
practices in Cambodia. Edible parts include bones and head (15). For spiders,
all parts were included as edible, following traditional consumption practice.
COMPLEMENTARY FOOD IN CAMBODIA
TABLE 2
Nutrient composition of intervention foods per 100 g dry weight1
Nutrient composition
WF2
WF-L
CSB++
CSB+3
Energy, kcal
Protein, g
Fat, g
Vitamin C, mg
Thiamin, mg
Riboflavin, mg
Niacin, mg
Vitamin B-6, mg
Folate, mg
Vitamin B-12, mg
Vitamin A, mg
Calcium,4 mg
Total iron,4 mg
Zinc,4 mg
474
15.4
10.3
0.2
0.2
0.1
5.2
0.4
12
2
35
570
4.2
4.5
428
12.6
9.2
100
0.1
0.5
4.8
1.7
60
2
166
631
6.3
5.2
458
16.8
10.7
100
0.1
0.5
4.8
1.7
60
2
166
277
10.5
7.0
482
14.6
16
100
0.1
0.5
4.8
1.7
60
2
166
173
9.9
6.6
1
Food groups: CSB+, CSB++, WF, WF-L. Values for macronutrients
and minerals were analyzed in samples of all 4 foods. Values for energy
contents were calculated from macronutrient contents. WF-L, CSB++, and
CSB+ were fortified with the same mineral and vitamin mix following the
specifications for CSB+ and CSB++ from World Food Programme specifications (version 1.1, 2010) (9, 10). CSB+, corn-soy blend plus; CSB++,
corn-soy blend plus plus; WF, WinFood; WF-L, WinFood-Lite.
2
The vitamin values for WF were calculated from food composition
values of 77% rice and 14% fish. The nutrient composition of rice and fish
was obtained from the USDA (rice: 50446; fish: 15089) (43).
3
Energy content estimation in CSB+ includes oil distributed to be added
at the time of preparation.
4
Values for calcium, iron, and zinc are analyzed values. WF-L, CSB+,
and CSB++ were added mineral premix, which should contribute 130 mg
calcium (added as mono- or dicalcium phosphate), 6.5 mg Fe (added as 4.0 mg
ferrous fumerate and 2.5 mg iron-sodium EDTA), and 5 mg zinc (added as zinc
oxide).
Randomization and blinding
Random allocation sequences were computer generated and
stratified by sex with varying block sizes of 12 and 24. All 4 products
were packed in sachets with an identical WinFood logo. Product
identification was single-blinded for investigators and enumerators
through the entire data collection until the preliminary analyses were
completed. The product identification (WF, WF-L, CSB+, or
CSB++) was marked in small print on the backside of each sachet,
allowing one staff member responsible for the food distribution to
ensure correct distribution of the products to each participant.
Complementary food products
The WF and WF-L products were locally produced by processing to a dry precooked semi-instant porridge. WF contained
small indigenous fish species (Esomus longimanus and Paralaubuca typus) and edible spiders (Haplopelma species) (Table
1). Fish species and edible spiders were selected among a range
of local foods based on screening for contents of iron and zinc.
The fish were small indigenous fish, which E. longimanus previously was identified to have a high iron content (15). The
selected spider, traditionally regarded as edible in Cambodia, is
traded in local food markets. Fish and spiders were procured
fresh directly from fishermen and traders by the project staff.
The fish in both products were initially sun-dried and protected
from contamination, and spiders were heat dried in an oven.
3 of 10
Dried fish and spiders were stored at 2208C until final processing.
The WF and WF-L were processed and packed at a medium-scale
food-processing facility (So! Nutritious Co. Ltd. in Phnom Penh)
under close supervision by the study team. Ingredients were
ground, mixed, and precooked by extrusion to obtain products
similar to the standard products from WFP with regard to texture
and cooking time. The final products were microbiologically
tested and approved for food safety at Institut Pasteur Cambodge
in Phnom Penh. The CSB products were donated by WFP and
produced outside Cambodia. The CSBs were repacked in sachets
of daily rations at So! Nutritious Co. Ltd. For CSB+, the specifications included adding 10% sugar and 10% oil by weight to the
product before cooking. Sugar was mixed into the CSB+ batch by
WFP before distribution. Cooking oil (sunflower oil) was received
from WFP and repacked in small, 5-g sachets and distributed
along with the CSB+ rations. Children aged 6–12 mo were given
one 5-g sachet of oil per day, while children aged 12–15 mo were
given 2 sachets per day.
Samples of the food products were analyzed for protein, fat, iron,
zinc, and calcium (Table 2). For CSB+, analyses were conducted on
the product with sugar added according to specifications but
without the additional oil, which was distributed separately. Protein
was analyzed by standard Kjeldahl nitrogen determination at the
University of Copenhagen, Denmark. Fat was analyzed by an
acidic Bligh and Dyer method (16) at Aarhus University, Denmark.
Iron, zinc, and calcium were analyzed by atomic absorption
spectrometry (Spektr-AA 200; Varian) after acidic digestion in
a DigiPREP MS Digestion System (SCP Science) (15).
Body composition
FFM and FM were assessed by using the deuterium dilution
technique to measure total body water following the protocol
developed by the International Atomic Energy Agency (17). The
infants were measured at baseline and endline. Each infant was
given an oral dose of 7 g deuterium oxide (99.8% 2 H 2 O)
(Cambridge Isotope Laboratories Inc.). The 2 H 2 O dose was
administered to the infant with a 10-mL syringe. Two preweighed paper towels were made available for each infant to
absorb any spilled 2H2O. One predose saliva sample was collected
before giving the 2H2O dose, and 2 postdose saliva samples were
collected 2 and 3 h, respectively, after the 2H2O dose was given.
Saliva samples were collected by putting a cotton ball in the
child’s mouth for 3–5 min. A thread was tied around the cotton
ball and kept hanging out of the mouth to prevent swallowing.
The wet cotton ball was removed from the child’s mouth and put
into a syringe, and the saliva was squeezed into a 1.5-mL cryotube.
Saliva samples were stored at 2208C until analysis for 2H2O
enrichment at St. Johns Research Institute, Bangalore, India.
Deuterium enrichment was measured by a Fourier transform infrared spectrometer (Shimadzu Corporation) and analyzed by the
software developed at the Dunn Unit in Cambridge, United
Kingdom. With data from the 3-h post sample, we calculated FFM
as total body water/hydration factor (17) by using sex- and agespecific hydration factors (18). FM was calculated as body weight
minus FFM (17). Before the study was unblinded, all infants with
FM ,5% were further reviewed and checked with field notes
regarding problems administrating the 2H2O to the child. Any
uncertainty of how much 2H2O was consumed by the child led to
his or her exclusion from the analyses.
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SKAU ET AL.
Iron status
Statistical analysis
Pediatric nurses took 3-mL samples of venous blood from
nonfasting subjects (venipuncture at left or right arm, supine
position) at baseline and at endline. A maximum of 2 attempts to
draw blood were set. Immediately after blood was collected,
blood drops were put on a hydrophobic glass slide for subsequent
blood hemoglobin measurement (2-fold analysis and control
samples) by using a HemoCue HB301 photometer (HemoCue).
Blood left in the syringe was filled into a heparin-coated
Vacutainer (Becton Dickinson) and kept chilled at 48C to prevent
microhemolysis. Then it was separated within 4 h by centrifugation (1300 3 g, 10 min at 48C). Plasma aliquots were kept
frozen at 2208C for 3 mo at baseline and endline, until they
were sent to the Department of Micronutrients, National Institute
of Nutrition, Hanoi, Vietnam, for further proceeding. Plasma ferritin, sTfR, a1-acid glycoprotein, and C-reactive protein concentrations were determined by enzyme immunoassays by using
commercial ELISA test kits (Ramco Laboratories). This procedure
was implemented according to protocols provided by the producers by using a BioTek Elx 808 microplate reader and BioTek
Gen5 v.1.07 software with 10% of samples analyzed in duplicates
and control samples. Controls that were used to check the precision and accuracy of each analytic method fell within the certified ranges for blood hemoglobin, plasma ferritin, sTfR, a1-acid
glycoprotein, and C-reactive protein. The between-assay CVs
were 0.5% for hemoglobin and ,10% for all ELISAs.
Data were double-entered in Epidata v.3.1 (The EpiData
Association) and analyzed by using STATA 12 for Windows
(StataCorp LP) and R 2.15 (The R Foundation). Plasma ferritin
concentrations were corrected for inflammation by using Creactive protein and a1-acid glycoprotein concentrations and the
correction factors as published by Thurnham and colleagues
(21). Baseline characteristics of the food groups were summarized by using descriptive statistics. ANOVA was used to estimate the mean change per group with corresponding 95% CIs.
Selected pairwise comparisons were considered: CSB+ was used
as the reference group, then CSB++. The primary analysis was
based on intention to treat, and secondary analysis was carried
out among children with high compliance with acquisition of
supplements, defined as missing no more than one of 9 food
distributions. A significance level of 5% was used.
Anthropometric measurements
All anthropometric measures were recorded at all time points
by the same 4 field assistants. All measurements except knee-heel
length were done in duplicate and the mean used in the analysis.
Weight was measured to the nearest 100 g by using an electronic
scale (SECA scale). Length was measured to the nearest 0.1 cm
(wooden board, borrowed from the WFP in Cambodia). Kneeheel length was measured as described previously (19) by using
a digital linear scale (Mitutoyo) with a resolution of 0.01 mm,
mounted with knee and heel caps cast in hard plastic. The result
was expressed as the mean of 5 consecutive separate measurements, calculated by the instrument. Skinfolds were measured by
a Harpenden caliper to the nearest 2 mm. MUAC and head
circumference were measured with nonstretchable plastic tapes
(Lasso-O tape; Harlow Printing Ltd.) to the nearest millimeter.
Anthropometric z scores were calculated based on WHO’s 2006
Child Growth Standards (20) by using Anthro v.3.1.
Additional data were obtained on morbidity, measured by selfreported illness history 2 wk before each monthly data collection
time point. Data on breastfeeding, introduction to complementary
food, and several sociodemographic variables were obtained at
baseline. Compliance was measured by a questionnaire on food
sharing and by sachet count: all daily ration sachets (both empty
and unopened) were returned and counted before the caretakers
could get a new monthly ration.
Sample size
Approximately 100 children per group were needed to detect
a difference of 0.4 SD, assuming 80% power and a 5% significance
level. To allow for 10% loss to follow-up, we aimed at recruiting
a total of 440 children.
RESULTS
Of the 514 invited infants from the study area, 440 were
screened. Of these, 419 met the inclusion criteria and were
randomly allocated to one of the 4 food groups (Figure 1). Of the
419 infants randomly allocated, 358 (85.4%) completed the
study. The main reason for loss to follow-up was migration due
to severe flooding of the area in August 2011.
Randomization resulted in baseline equivalence (Table 3),
although the CSB++ group had a slightly higher proportion of
children with LAZ ,22.0 at baseline. Almost all children were
currently breastfed, and the mean age at introduction to complementary food was 5.3 mo. No difference was found between
the 4 groups in the prevalence of several morbidity symptoms at
each time point (data not shown).
During the 9-mo intervention period, mean weight increased
by 1.73 kg (95% CI: 1.68, 1.78), from 6.85 to 8.58 kg. The weight
increase was due to a 1.96-kg (95% CI: 1.88, 2.03) increment in
mean FFM and a 0.21-kg (95% CI: 0.15, 0.28) decline in FM. No
differences were found between any of the WinFood products and
CSB+ or CSB++ with respect to changes in FFM and FM (Table
4). Similarly, no differences were found in the change in plasma
ferritin, sTfR, and hemoglobin concentration between any of the
WinFood products and the CSB+ or CSB++. Plasma ferritin
concentrations decreased and plasma sTfR concentrations increased over the intervention period in all food groups (P , 0.01
for both), indicating a deterioration of iron status in all infants
(Table 4). The WF group had a higher prevalence of anemia
(53.7%) at endline compared with the other 3 intervention
groups (35.2%, 35.2%, and 39.8%, respectively, for CSB+,
CSB++, and WF-L; P , 0.05). Based on counts of empty
sachets, 57.8%, 62.5%, 56.0%, and 55.2% of the distributed WF,
WF-L, CSB+, and CSB++, respectively, were consumed. At all
follow-up visits, all caregivers reported that the distributed foods
had been eaten only by the infant and not shared with the
household. A secondary analysis was conducted among the 288
(68.7%) with high compliance (Table 5). Based on counts of
empty sachets in this group, 75.3%, 75.9%, 73.8%, and 75.3%,
respectively, of the distributed WF, WF-L, CSB++, and CSB+
were consumed. No differences in FFM and FM or plasma
ferritin and sTfR were found between the intervention and the
reference products. The WF group had a marginally lower
change in hemoglobin than CSB++ (Table 5).
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COMPLEMENTARY FOOD IN CAMBODIA
FIGURE 1 Flow diagram of the study participants. Food groups: CSB+, corn-soy blend plus; CSB++, corn-soy blend plus plus; WF, WinFood; WF-L,
WinFood-Lite; WLZ, weight-for-length z score. Participants missing a maximum of one food distribution are viewed as having high compliance.
For total weight, we found that the WF group had a smaller, but
not significant, increase compared with the CSB++ group (Table
6). The larger increase in knee-heel length in the WF-L group
compared with the CSB+ reference group was not significant.
Also, the reduced growth in knee-heel length in the WF group
compared with the CSB++ group was not significant (Table 6).
Moreover, no differences were found between the WinFood
products and CSB+ or CSB++ in MUAC, skinfolds, and head
circumference (Supplemental Tables 1 and 2).
To assess the growth over time, we plotted the changes in mean
anthropometric indices as a function of the monthly follow-up
visits (Figure 2). For LAZ and WAZ, the patterns indicated that
WF and CSB+ had an earlier decline compared with WF-L and
CSB++. WLZ showed similar patterns in the 4 groups over time.
No difference was found in the mean change of z scores over
time between WF-L and any of the reference groups on any of
the indices. WF was not different from CSB+ with respect to any
of the anthropometric indices. However, compared with CSB++,
WF was associated with 20.20 LAZ (95% CI: 20.39, 20.01)
and 20.25 WAZ (95% CI: 20.03, 20.46).
In a secondary analysis, the 2 CSB products were compared
with each other, and the 2 WinFood products were compared with
TABLE 3
Baseline characteristics of study participants by food group1
Characteristic
WF
WF-L
CSB++
CSB+
Children, n
Child characteristics
Sex, boys, n (%)
Child age, mo
Currently breastfeeding, n (%)
Exclusive breastfeeding, n (%)
Age introduced to solid food, mo
Weight, kg
Length, cm
Hemoglobin, g/L
Weight-for-length z score
Children with z score ,22, n (%)
Length-for-age z score
Children with z score ,22, n (%)
Weight-for-age z score
Children with z score ,22, n (%)
Maternal characteristics
Midupper arm circumference, cm
Schooling, y
Household characteristics
Total household members
Children aged ,5 y
Access to protected well, n (%)
Flush toilet or pit latrine, n (%)
Primary income, n (%)
Fishery
Farming
106
104
103
106
56
5.8
102
71
5.2
6.9
64.5
108
20.30
2
20.89
13
20.85
12
(52.8)
6 0.5
(96.2)
(67.0)
6 1.5
6 0.9
6 2.5
69
6 0.99
(1.9)
6 0.99
(12.3)
6 1.00
(11.3)
24.7 6 2.8
562
5.4
1.3
61
25
6 2.0
6 1.0
(57.5)
(23.6)
3 (2.8)
60 (56.6)
54
5.9
101
70
5.3
6.9
64.6
107
20.37
4
20.90
13
20.89
14
(51.9)
6 0.6
(97.1)
(67.3)
6 1.4
6 0.8
6 2.3
69
6 0.92
(3.9)
6 0.97
(12.5)
6 0.95
(13.5)
24.9 6 2.6
563
5.1
1.3
58
26
6 1.0
6 1.0
(55.8)
(25.0)
2 (1.9)
63 (60.6)
54
5.9
101
73
5.3
6.8
64.1
108
20.39
7
21.12
22
21.06
21
(52.4)
6 0.6
(98.1)
(70.9)
6 1.5
6 0.9
6 2.7
69
6 1.03
(6.8)
6 1.05
(21.4)
6 1.10
(20.4)
24.3 6 2.8
562
5.3
1.4
59
22
6 2.0
6 1.0
(57.3)
(21.4)
6 (5.8)
66 (64.1)
56
5.9
106
73
5.3
6.8
64.4
108
20.41
5
20.96
14
20.97
12
(52.8)
6 0.6
(100)
(68.9)
6 1.5
6 0.8
6 2.4
69
6 0.90
(4.7)
6 0.88
(13.2)
6 0.90
(11.3)
24.3 6 2.5
662
5.5
1.4
54
19
6 2.0
6 1.0
(50.9)
(17.9)
6 (5.7)
63 (59.4)
1
Data are means 6 SDs, unless stated otherwise. Food groups: CSB+, corn-soy blend plus; CSB++, corn-soy blend
plus plus; WF, WinFood; WF-L, WinFood-Lite.
6 of 10
SKAU ET AL.
TABLE 4
Primary outcome for all participants: effects of WF and WF-L compared with “standard product” (CSB+) and the “best product” (CSB++) after a 9-mo
intervention on body composition and iron status1
Characteristic
Fat-free mass, kg
Plasma
ferritin, mg/L
Plasma transferrin
receptor, mg/L
42.1
41.5
47.3
46.5
(30.9,
(33.6,
(33.4,
(34.7,
(97)
(95)
(94)
(95)
3.6
3.7
3.6
3.7
(3.4,
(3.5,
(3.4,
(3.5,
3.8)
3.9)
3.8)
4.0)
(101)
(101)
(99)
(101)
107
106
108
107
(105,
(104,
(106,
(105,
109)
108)
109)
109)
(104)
(103)
(100)
(104)
11.9
13.7
12.6
11.9
(9.6, 14.1) (83)
(11.3, 16.1) (87)
(10.1, 15.1) (86)
(10.0, 13.9) (87)
9.9
9.3
9.2
9.3
(9.0,
(8.6,
(8.4,
(8.5,
10.8) (83)
10.0) (88)
9.9) (86)
10.0) (87)
105
107
108
107
(102,
(104,
(105,
(104,
107)
108)
110)
109)
(82)
(88)
(87)
(88)
Fat mass, kg
Age, 6 mo
WF
5.34 (5.21, 5.47) (103)
1.54 (1.45, 1.63) (103)
WF-L
5.40 (5.27, 5.52) (96)
1.52 (1.42, 1.63) (96)
CSB++
5.29 (5.16, 5.42) (93)
1.50 (1.39, 1.60) (93)
CSB+
5.39 (5.27, 5.51) (98)
1.44 (1.34, 1.53) (98)
Age, 15 mo
WF
7.25 (7.05, 7.46) (70)
1.27 (1.16, 1.38) (70)
WF-L
7.40 (7.20, 7.59) (71)
1.34 (1.22, 1.47) (71)
CSB++
7.33 (7.14, 7.53) (74)
1.24 (1.09, 1.34) (74)
CSB+
7.32 (7.16, 7.49) (78)
1.25 (1.15, 1.36) (78)
Difference (15–6 mo)
compared with
standard product
(CSB+)2
WF
0.04 (20.20, 0.28) (67) 20.11 (20.34, 20.11) (67)
WF-L
0.14 (20.10, 0.38) (66) 20.03 (20.26, 0.19) (66)
Best product
(CSB++)3
WF
20.03 (20.27, 0.21) (67) 20.05 (20.28, 0.17) (67)
WF-L
0.07 (20.18, 0.31) (66)
0.03 (20.20, 0.25) (66)
53.3)
49.4)
61.2)
58.4)
Hemoglobin, g/L
2.1 (219.9, 24.1) (75)
0.0 (21.1, 1.2) (79) 20.2 (20.6, 0.2) (80)
6.7 (214.9, 28.3) (80) 20.4 (21.6, 0.7) (86)
0.2 (20.1, 0.5) (88)
2.5 (219.5, 24.6) (75)
7.1 (214.6, 28.8) (80)
0.5 (20.7, 1.6) (79) 20.3 (20.7, 0.1) (80)
0.0 (21.1, 1.1) (86)
0.1 (20.3, 0.5) (88)
1
Data were analyzed by the intention-to-treat approach and are presented as mean differences; 95% CIs in parentheses (n). Food groups: CSB+, corn-soy
blend plus; CSB++, corn-soy blend plus plus; WF, WinFood; WF-L, WinFood-Lite.
2
Standard corn-soya blend product.
3
Corn-soya blend product improved for infants and young children.
each other. The increase in knee-heel length was 1.9 mm (95%
CI: 0.1, 3.8) in the CSB++ group compared with the CSB+ group.
When only including participants with high compliance, the CSB++
group showed an increase of 2.4 mm (95% CI: 0.3, 4.5) in kneeheel length and also an increase of 0.5 cm (95% CI: 0.01, 1.0) in
total length compared with the CSB+ group. Moreover, we found
TABLE 5
Primary outcome for participants with high compliance: effects of WF and WF-L compared with “standard product” (CSB+) and the “best product” (CSB++)
after a 9-mo intervention on body composition and iron status1
Characteristic
Age, 6 mo
WF
WF-L
CSB++
CSB+
Age, 15 mo
WF
WF-L
CSB++
CSB+
Difference (15–6 mo)
compared with
standard product
(CSB+)2
WF
WF-L
Best product
(CSB++)3
WF
WF-L
Fat-free mass, kg
Fat mass, kg
Plasma
ferritin, mg/L
Plasma transferrin
receptor, mg/L
5.31
5.31
5.29
5.42
(5.16,
(5.16,
(5.14,
(5.27,
5.46)
5.46)
5.44)
5.58)
(68)
(68)
(62)
(63)
1.52
1.57
1.49
1.47
(1.41,
(1.43,
(1.37,
(1.35,
1.64)
1.70)
1.61)
1.59)
(68)
(70)
(62)
(63)
46.0
39.4
51.5
46.0
(30.1,
(30.5,
(31.9,
(31.5,
61.9)
48.4)
71.1)
60.6)
(64)
(72)
(64)
(62)
3.6
3.8
3.6
3.7
(3.3,
(3.5,
(3.3,
(3.3,
3.9)
4.1)
3.8)
3.9)
(66)
(77)
(69)
(66)
7.21
7.43
7.33
7.36
(6.98,
(7.22,
(7.10,
(7.18,
7.43)
7.64)
7.55)
7.55)
(60)
(63)
(60)
(62)
1.27
1.34
1.27
1.25
(1.15,
(1.20,
(1.14,
(1.14,
1.38)
1.48)
1.41)
1.36)
(60)
(63)
(60)
(62)
12.0
13.4
11.6
11.9
(9.6, 14.5) (70)
(10.7, 16.2) (72)
(9.5, 13.6) (70)
(9.8, 13.9) (68)
9.9
9.1
8.8
9.3
(8.9,
(8.4,
(8.0,
(8.4,
11.0) (70)
9.9) (73)
9.5) (70)
10.2) (68)
Hemoglobin, g/L
107
107
108
107
(105,
(104,
(106,
(105,
110)
109)
109)
110)
(69)
(77)
(69)
(68)
104
107
108
107
(101,
(105,
(106,
(104,
107)
109)
111)
109)
(69)
(73)
(71)
(69)
0.00 (20.26, 0.27) (58)
0.16 (20.10, 0.42) (58)
20.10 (20.34, 0.15) (58)
20.02 (20.26, 0.23) (58)
2.1 (223.0, 27.2) (63)
7.5 (217.2, 32.3) (67)
0.0 (21.3, 1.3) (66)
20.6 (21.9, 0.7) (73)
20.3 (20.7, 0.2) (68)
0.2 (20.3, 0.6) (73)
20.03 (20.30, 0.23) (58)
0.12 (20.15, 0.38) (58)
20.15 (20.40, 0.01) (58)
20.07 (20.32, 0.18) (58)
6.9 (218.0, 31.8) (63)
12.4 (212.2, 36.9) (67)
0.6 (20.7, 1.9) (66)
0.1 (21.2, 1.3) (73)
20.4 (20.9, 0.0) (68)
0.0 (20.4, 0.5) (73)
1
Participants included those who missed at most one food distribution. Data are presented as mean differences; 95% CIs in parentheses (n). Food groups:
CSB+, corn-soy blend plus; CSB++, corn-soy blend plus plus; WF, WinFood; WF-L, WinFood-Lite.
2
Standard corn-soya blend product.
3
Corn-soya blend product improved for infants and young children.
7 of 10
COMPLEMENTARY FOOD IN CAMBODIA
TABLE 6
Secondary outcomes for all participants and for participants with high compliance: effect of WF and WF-L compared
with the “standard product” (CSB+) and the “best product” (CSB++) after a 9-mo intervention on weight, length, and
knee-heel length1
Characteristic
Weight, kg
Length, cm
All participants
Age, 6 mo
WF
6.9 (6.7, 7.1) (106)
WF-L
6.9 (6.7, 7.1) (104)
CSB++
6.8 (6.6, 6.9) (103)
CSB+
6.9 (6.7, 7.0) (106)
Age, 15 mo
WF
8.5 (8.3, 8.7) (85)
WF-L
8.7 (8.5, 8.9) (93)
CSB++
8.5 (8.3, 8.7) (88)
CSB+
8.5 (8.4, 8.7) (92)
Difference (15–6 mo) compared with
standard product (CSB+)2
WF
0.0 (20.2, 0.1) (85)
WF-L
0.1 (20.1, 0.2) (93)
Best product (CSB++)3
WF
20.1 (20.3, 0.1) (85)
WF-L
0.0 (20.2, 0.2) (93)
All participants with high compliance4
Age, 6 mo
WF
6.8 (6.6, 7.0) (70)
WF-L
6.9 (6.7, 7.1) (77)
CSB++
6.7 (6.5, 6.9) (71)
CSB+
6.9 (6.7, 7.1) (70)
Age, 15 mo
WF
8.5 (8.34, 8.7) (70)
WF-L
8.8 (8.5, 9.0) (77)
CSB++
8.6 (8.4, 8.8) (71)
CSB+
8.6 (8.4, 8.8) (70)
Difference (15–6 mo) compared with
standard product (CSB+)2
WF
20.1 (20.2, 0.1) (70)
WF-L
0.1 (20.04, 0.3) (77)
Best product (CSB++)3
WF
20.2 (20.4, 20.01) (70)5
WF-L
0.0 (20.2, 0.2) (77)
Knee-heel length, mm
64.5
64.6
64.1
64.4
(64.0,
(64.1,
(63.6,
(64.0,
65.1)
65.1)
64.7)
65.1)
(106)
(104)
(103)
(106)
168.9
170.1
169.3
170.3
(167.1,
(168.6,
(167.6,
(168.9,
170.7)
171.5)
171.0)
171.7)
(105)
(104)
(103)
(104)
75.0
75.2
74.9
74.7
(74.4,
(74.7,
(74.3,
(74.2,
75.6)
75.7)
75.5)
75.2)
(85)
(93)
(88)
(92)
203.9
206.2
205.6
204.7
(201.9,
(204.4,
(203.7,
(203.0,
206.0)
208.0)
207.6)
206.4)
(85)
(93)
(88)
(92)
0.2 (20.3, 0.6) (85)
0.3 (20.2, 0.7) (93)
0.6 (21.3, 2.5) (84)
1.7 (20.1, 3.5) (92)
20.2 (20.7, 0.3) (85)
20.1 (20.6, 0.3) (93)
21.3 (23.2, 0.5) (84)
20.2 (22.1, 1.6) (92)
64.4
64.7
64.1
64.2
(63.8,
(64.2,
(63.5,
(63.6,
65.0)
65.2)
64.7)
64.8)
(70)
(77)
(71)
(70)
168.6
169.9
169.0
170.2
(166.6,
(168.3,
(167.0,
(168.6,
170.7)
171.5)
171.0)
171.9)
(69)
(76)
(71)
(69)
74.9
75.4
75.0
74.6
(74.2,
(74.8,
(74.3,
(74.0,
75.6)
75.9)
75.7)
75.2)
(70)
(77)
(71)
(70)
203.9
206.4
206.0
205.1
(201.5,
(204.5,
(203.8,
(203.1,
206.4)
208.4)
208.2)
207.2)
(70)
(77)
(71)
(70)
0.1 (20.4, 0.6) (70)
0.3 (20.2, 0.8) (77)
0.4 (21.7, 2.5) (69)
1.8 (20.2, 3.9) (76)
20.4 (20.9, 0.1) (70)
20.2 (20.7, 0.3) (77)
22.0 (24.0, 0.1) (69)
20.6 (22.6, 1.5) (76)
1
Data were analyzed by the intention-to-treat approach and are presented as mean differences; 95% CIs in parentheses
(n). Participants included those who missed at most one food distribution. Food groups: CSB+, corn-soy blend plus;
CSB++, corn-soy blend plus plus; WF, WinFood; WF-L, WinFood-Lite.
2
Standard corn-soya blend product.
3
Corn-soya blend product improved for infants and young children.
4
Participants who missed at most 1 food distribution.
5
No significant difference after multiplicity adjustment of the P value (using Bonferroni).
that the difference between CSB++ and CSB+ in length growth
was 0.19 LAZ (95% CI: 0.00, 0.39), whereas the difference
between WF-L and WF in weight was 0.20 WAZ (95% CI:
20.01, 0.42).
DISCUSSION
No differences were found in our primary outcomes, the increments of FFM and change in iron status, between WF and WFL than either of the 2 CSB products. Also, the secondary outcome
measurements for growth did not show significant differences
between WF and WF-L compared with either of the 2 CSB
products. For iron status, it was discouraging to find that, although all groups received improved complementary food of
which 3 products were iron fortified for 9 mo, iron status de-
teriorated. On the basis of dietary data from the same population,
we recently modeled the nutritious sufficiency of the diets of 6- to
12-mo-old Cambodian infants by using linear programming. The
modeling indicated that the 4 intervention foods supplied insufficient iron to meet the iron needs for the infants (22). This
indication from the modeling is now supported by biochemical
evidence for iron status from the present study.
No significant differences were found in the FFM point estimates, but it is worth noticing that the highest increment was
seen in WF-L. For the subgroup of infants with high compliance
to the food products, the increment was 0.16 kg (95% CI: 20.1,
0.42) higher in the WF-L group compared with the CSB+ group,
equivalent to an 8% higher increment in FFM over the 9-mo
intervention. Comparing the body composition data with reference
data based on healthy infants from the United States (23) (no
8 of 10
SKAU ET AL.
FIGURE 2 Mean change in anthropometric indices from baseline at each monthly follow-up visit in participants with high compliance (WF: n = 55–70;
WF-L: n = 66–77; CSB++: n = 53–71; CSB+: n = 56–70). (A) Mean changes in LAZ, (B) mean changes in WAZ, and (C) mean changes in WLZ. The mean
changes between baseline and endpoint were analyzed by ANOVA, adjusted for multiplicity. a,bDifferences of P , 0.05. CSB+, corn-soy blend plus; CSB++,
corn-soy blend plus plus; LAZ, length-for-age z score; WAZ, weight-for-age z score; WF, WinFood; WF-L, WinFood-Lite; WLZ, weight-for-length
z score.
COMPLEMENTARY FOOD IN CAMBODIA
Cambodian reference data are available), the infants in the present
study at 6 mo of age had similar FFM (5.3 vs. 5.4 kg) but lower FM
(1.5 vs. 2.4 kg). At endline, all groups had a lower FFM (;0.4 kg)
and FM (;1.4 kg) compared with this reference group (calculated
by interpolating data from 12 to 18 mo). Thus, body fat in this
Cambodian population is low compared with healthy American
children, and an intervention with a daily supplement of nutritious
complementary food was not able to increase fat deposition.
The secondary anthropometric outcomes did not show differences between the food groups. However, the measurement of
growth in knee-heel length was indicatively highly sensitive to
marginal impacts of the intervention foods on length growth.
Because stunting is particularly difficult to prevent in food-insecure populations, the finding has relevance for improving the
monitoring of impact of nutrition programs on stunting. There are
several explanations for the possible higher sensitivity of the
knee-heel length compared with total length. First, the precision
of measuring knee-heel length is higher than measuring total
length (19). The error of one measuring sequence was 0.55 mm,
equal to ;3 d of growth in this population (unpublished data).
Second, the increments in lower leg length are relatively higher
than increments in the remaining length of the body. Lower leg
length contributes ;25% of total length at birth and ;29% at
5 y (24). In the present study, the percentages were 26.3%
at baseline and 27.4% at the end of the study, whereas 33.5% of
the increment in total length during the intervention was due to
increments of knee-heel length. Some data suggest that dairy
intake has a specific effect on leg growth (25), but we do not
know if this is the case for other ASFs such as fish. Interestingly,
a number of studies have reported beneficial associations between leg length and adult health (26, 27).
Studies assessing the impact of improved locally produced
complementary foods on growth are highly diverse in the type of
intervention products and study designs (28–35). Three studies
assessing complementary foods containing ASF, by comparing to
foods with micronutrient fortification or lipid-based fortified
spread, showed no clear effect on linear growth (28–30). A study
from Malawi showed that CSB++ was not inferior to support linear
growth compared with lipid-based ready-to-use supplementary
foods (36). In the present study, infants given WF-L had a growth
pattern over the intervention period similar to that of CSB++, with
a later and less marked decline in LAZ and WAZ compared with
CSB+. WF showed a growth pattern over time in LAZ and WAZ,
which did not differ from CSB+. Explanations for these apparent
differences in growth patterns can be sought in the nutritional
composition of the foods. WF-L and CSB++ differ from the 2 other
products by containing a component of ASF and being fortified.
Fish are the most accessible ASF in many low-income
countries. In general, the iron content of fish is less than that of
red meat but similar to the content in chicken and pork (37), but
small fish, in which most tissues are edible, are in general better
iron sources than larger fish, in which only the fillet is eaten (38).
Furthermore, small fish eaten with bones are a good calcium
source, with a calcium bioavailability similar to milk (39). In
Cambodia, the 2013 price of dried fish powder was about 1.7 US
$/kg, corresponding to around 2.8 US$/kg ASF protein, whereas
the price of skimmed milk powder was 3.7 US$/kg (40), or more
than 7 US$/kg ASF protein, because lactose contributes about
half of skimmed milk powder. Hence, the cost aspect is an incentive for further exploration of using small fishes in processed
9 of 10
complementary food as an alternative to milk powder to enhance
the nutritional quality of complementary feeding in food-insecure
settings with access to aquatic resources. WF had the highest
proportion of ASF from a combination of fish (E. longimanus and
P. typus) and edible spiders (Haplopelma species). The Haplopelma species was selected because of a high content of zinc
(16 mg zinc/100 g raw weight; unpublished data). There is
a growing interest in using edible insects as an alternative protein source for human consumption, and studies indicate that
insects are good sources of micronutrients (41, 42). However, to
our knowledge, there are no studies of the bioavailability of
minerals or other nutrients in humans from edible insects, and
studies on nutrient composition and bioavailability from edible
insects are needed.
The study zone experienced heavy flooding during the intervention period, and this may have introduced a higher dropout
rate than expected. Many of the households in flooded areas faced
a serious challenge in reaching the distribution sites, and the
study team made particular efforts to reach these households.
However, the high number of dropouts has weakened the power
of the study. Most caregivers said they did not share the supplemental food in the household, and we cannot preclude that food
sharing occurred. However, the packaging into very small individual sachets made food sharing less tempting and more
cumbersome. If the child did not eat the whole portion, food could
have been lost as plate waste. The pace of transition from breast
milk to complementary food may vary between the infants, and
complete compliance to predefined portion sizes was not expected.
Overall, the locally produced product did not differ from the
CSBs products. The results indicate that supplementary food
products distributed for complementary feeding in food-insecure
populations benefit from being fortified and containing ASF. In
Cambodia, small fishes have potential as a cheap and sustainable
local ASF source that is an alternative to milk, which can contribute to improve the nutritional quality of locally processed
fortified complementary foods and food aid products.
We thank Dr. Chan Theary and Dr. Chan Ketsana from the Reproductive
and Child Health Alliance (RACHA) for helping to implement the study in
PeaRieng, Prey Veng, and assisting with facilitating contact with the mothers.
We also thank Dr. Hout Kalyan and Dr. Seng Narin from the PeaRieng referral
hospital for support during data collection. We thank Sok Seyha, Lach Thea,
Ao Veasna, Ann Kim Eng, Tech Sivyong, Em Thearith, Dy Moeunnary, Sok
Daream, and Khov Kuong, staff at the Department of Fisheries Post-harvest
Technologies and Quality Control, Fishery Administration, Ministry of Agriculture, Forestry and Fisheries, Cambodia, for their great commitment at field
works, as well as Graham Taylor and Marjorie Negado from So! Nutritious Co.
Ltd. for helping with the production of WF and WF-L.
The authors’ responsibilities were as follows—JKHS, TB, CC, CM, SF,
FTW, MAD, JB, HF, KFM, and NR: designed the study; JKHS, TB, CC,
CM, and JM: conducted the study; JKHS, TB, CC, FTW, MAD, JB, and NR:
developed the WinFood products; USU: conducted the Fourier transform
infrared spectrometer analyses; JKHS, JCW, CR, HF, KFM, and NR: analyzed
the data; JKHS: wrote the first draft of the manuscript. All authors edited the
manuscript and approved the final version. None of the authors reported any
conflicts of interest related to this study.
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