ABSTRACT

OBJECTIVE:

This study aimed to describe the prevalence and predictors of childhood anemia in an Amazonian population-based birth cohort study.

METHODS:

Prevalence of maternal anemia was estimated at delivery (hemoglobin [Hb] concentration < 110 g/L) in women participating in the MINA-Brazil birth cohort study and in their children, examined at ages one, two (Hb < 110 g/L), and five (Hb < 115 g/L). Moreover, ferritin, soluble transferrin receptor, and C-reactive protein concentrations were measured in mothers at delivery and in their 1- and 2-year-old children to estimate the prevalence of iron deficiency and its contribution to anemia, while adjusting for potential confounders by multiple Poisson regression analysis (adjusted relative risk [RR_a]).

RESULTS:

The prevalence 95% confidence interval (CI) of maternal anemia, iron deficiency, and iron-deficiency anemia at delivery were 17.3% (14.0–21.0%), 42.6% (38.0–47.2%), and 8.7% (6.3–11.6)%, respectively (n = 462). At one year of age (n = 646), 42.2% (38.7–45.8%) of the study children were anemic, 38.4% (34.6–42.3%) were iron-deficient, and 26.3 (23.0–29.9) had iron-deficiency anemia. At two years of age (n = 761), these values decreased to 12.8% (10.6–15.2%), 18.1% (15.5–21.1%), and 4.1% (2.8–5.7%), respectively; at five years of age (n = 655), 5.2% (3.6–7.2%) were anemic. Iron deficiency (RR_a = 2.19; 95%CI: 1.84–2.60) and consumption of ultra-processed foods (UPF) (RR_a = 1.56; 95%CI: 1.14–2.13) were significant contributors to anemia at 1 year, after adjusting for maternal schooling. At 2 years, anemia was significantly associated with maternal anemia at delivery (RR_a: 1.67; 95%CI: 1.17–2.39), malaria since birth (2.25; 1.30–3.87), and iron deficiency (2.15; 1.47–3.15), after adjusting for children's age and household wealth index.

CONCLUSIONS:

Anemia continues to be highly prevalent during pregnancy and early childhood in the Amazon. Public health policies should address iron deficiency, UPF intake, maternal anemia, and malaria to prevent and treat anemia in Amazonian children.

DESCRIPTORS:
Anemia, Iron Deficiency; Malaria; Risk Factors; Cohort Studies

INTRODUCTION

Nearly one-third of the world's population is estimated to be anemic, with increased morbidity and mortality, decreased work productivity, and impaired child neurodevelopment¹1 Chaparro CM, Suchdev PS. Anemia epidemiology, pathophysiology, and etiology in low- and middle-income countries. Ann N Y Acad Sci. 2019 Aug;1450(1):15-31. https://doi.org/10.1111/nyas.14092
https://doi.org/10.1111/nyas.14092... . Infants, young children, and pregnant women are among the populations most vulnerable to anemia worldwide²2 Hess SY, Owais A, Jefferds ME, Young MF, Cahill A, Rogers LM. Accelerating action to reduce anemia: review of causes and risk factors and related data needs. Ann N Y Acad Sci. 2023 May;1523(1):11-23. https://doi.org/10.1111/nyas.14985
https://doi.org/10.1111/nyas.14985... . From 2000 to 2019, the prevalence of anemia in children aged 6–59 months decreased from 48% to 40% worldwide and from 30% to 20% in Latin America and the Caribbean; however, the burden of childhood anemia remains staggering in many low- and middle-income countries³3 Stevens GA, Paciorek CJ, Flores-Urrutia MC, Borghi E, Namaste S, Wirth JP, et al. National, regional, and global estimates of anaemia by severity in women and children for 2000-19: a pooled analysis of population-representative data. Lancet Glob Health. 2022 May;10(5):e627-39. https://doi.org/10.1016/S2214-109X(22)00084-5
https://doi.org/10.1016/S2214-109X(22)00... . Recent meta-analyses reveal regional disparities in the prevalence of childhood anemia in Brazil that are hidden behind pooled countrywide estimates⁴4 Ferreira HS, Vieira RC, Livramento AR, Dourado BL, Silva GF, Calheiros MS. Prevalence of anaemia in Brazilian children in different epidemiological scenarios: an updated meta-analysis. Public Health Nutr. 2021 Jun;24(8):2171-84. https://doi.org/10.1017/S1368980019005287
https://doi.org/10.1017/S136898001900528... ,⁵5 Nogueira-de-Almeida CA, Ued FD, Del Ciampo LA, Martinez EZ, Ferraz IS, Contini AA, et al. Prevalence of childhood anaemia in Brazil: still a serious health problem: a systematic review and meta-analysis. Public Health Nutr. 2021 Dec;24(18):6450-65. https://doi.org/10.1017/S136898002100286X
https://doi.org/10.1017/S136898002100286... . For example, the Brazilian National Survey on Child Nutrition (ENANI-2019) estimated that 10% of the preschool children living in Brazilian metropolitan areas were anemic; however, in the North—the region that comprises most of the Amazon Basin of Brazil—, the prevalence reached 17%⁶6 Castro IR, Normando P, Farias DR, Berti TL, Schincaglia RM, Andrade PG, et al. Factors associated with anemia and vitamin A deficiency in Brazilian children under 5 years old: Brazilian National Survey on Child Nutrition (ENANI-2019). Cad Saude Publica. 2023 Sep;25(39 Suppl 2):e00194922. https://doi.org/10.1590/0102-311xen194922
https://doi.org/10.1590/0102-311xen19492... . The corresponding estimates for children aged 6–23 months were 20% countrywide and 30% in the North⁶6 Castro IR, Normando P, Farias DR, Berti TL, Schincaglia RM, Andrade PG, et al. Factors associated with anemia and vitamin A deficiency in Brazilian children under 5 years old: Brazilian National Survey on Child Nutrition (ENANI-2019). Cad Saude Publica. 2023 Sep;25(39 Suppl 2):e00194922. https://doi.org/10.1590/0102-311xen194922
https://doi.org/10.1590/0102-311xen19492... .

Iron deficiency accounts for nearly 50% of the global burden of childhood anemia⁷7 Victora CG, Christian P, Vidaletti LP, Gatica-Domínguez G, Menon P, Black RE. Revisiting maternal and child undernutrition in low-income and middle-income countries: variable progress towards an unfinished agenda. Lancet. 2021 Apr;397(10282):1388-99. https://doi.org/10.1016/S0140-6736(21)00394-9
https://doi.org/10.1016/S0140-6736(21)00... ,⁸8 Brown KH, Moore SE, Hess SY, McDonald CM, Jones KS, Meadows SR, et al. Increasing the availability and utilization of reliable data on population micronutrient (MN) status globally: the MN Data Generation Initiative. Am J Clin Nutr. 2021 Sep;114(3):862-70. https://doi.org/10.1093/ajcn/nqab173
https://doi.org/10.1093/ajcn/nqab173... . For the primary prevention of iron-deficiency anemia in young children, the World Health Organization (WHO) recommends exclusive breastfeeding (EBF) for six months and continued breastfeeding (BF) until 2 years or older, along with adequate complementary feeding with iron or multiple micronutrient supplements⁹9 World Health Organization. Global strategy for infant and young child feeding. Geneva: World Health Organization; 2003 [cited 2023 May 22]. Available from: https://www.who.int/publications/i/item/9241562218
https://www.who.int/publications/i/item/... . However, supplementation programs to prevent anemia have a relatively low coverage in most low- and middle-income countries⁷7 Victora CG, Christian P, Vidaletti LP, Gatica-Domínguez G, Menon P, Black RE. Revisiting maternal and child undernutrition in low-income and middle-income countries: variable progress towards an unfinished agenda. Lancet. 2021 Apr;397(10282):1388-99. https://doi.org/10.1016/S0140-6736(21)00394-9
https://doi.org/10.1016/S0140-6736(21)00... and fail to address non-nutritional causes of childhood anemia, such as soil-transmitted helminthic infections, schistosomiasis, and malaria²2 Hess SY, Owais A, Jefferds ME, Young MF, Cahill A, Rogers LM. Accelerating action to reduce anemia: review of causes and risk factors and related data needs. Ann N Y Acad Sci. 2023 May;1523(1):11-23. https://doi.org/10.1111/nyas.14985
https://doi.org/10.1111/nyas.14985... .

This study focused on childhood anemia in the North of Brazil, the region with the highest prevalence of anemia in preschool children estimated in the most recent countrywide survey⁶6 Castro IR, Normando P, Farias DR, Berti TL, Schincaglia RM, Andrade PG, et al. Factors associated with anemia and vitamin A deficiency in Brazilian children under 5 years old: Brazilian National Survey on Child Nutrition (ENANI-2019). Cad Saude Publica. 2023 Sep;25(39 Suppl 2):e00194922. https://doi.org/10.1590/0102-311xen194922
https://doi.org/10.1590/0102-311xen19492... . We described the prevalence of anemia during the first five years of follow-up of children participating in an ongoing population-based birth cohort study in the Amazon and identified independent predictors of anemia risk that may constitute potential targets for public health interventions.

METHODS

Study Design, Population, and Data Collection

The Maternal and Child Health and Nutrition in Acre, Brazil (MINA-Brazil) birth cohort study started in 2015 with 1,246 mother-child pairs in the Western Amazonian city of Cruzeiro do Sul (CZS), Acre State. It aimed to characterize the effect of a wide range of early exposures on child health¹⁰10 Cardoso MA, Matijasevich A, Malta MB, Lourenco BH, Gimeno SG, Ferreira MU, et al.; MINA-Brazil Study Group. Cohort profile: the maternal and child health and nutrition in Acre, Brazil, birth cohort study (MINA-Brazil). BMJ Open. 2020 Feb;10(2):e034513. https://doi.org/10.1136/bmjopen-2019-034513
https://doi.org/10.1136/bmjopen-2019-034... . The infant mortality rate in CZS was estimated at 10.8 deaths per 1,000 live births in 2015 and 10.6 per 1,000 in 2020¹¹11 Instituto Brasileiro de Geografia e Estatística. Cities and states. Brazil. Rio de Janeiro: Instituto Brasileiro de Geografia e Estatística; 2022 [cited 2023 May 22]. Available from: https://ibge.gov.br/cidades-e-estados/ac/cruzeiro-do-sul.html
https://ibge.gov.br/cidades-e-estados/ac... . CZS experiences year-round malaria transmission, with nearly 90% of all malarial infections in young children being due to Plasmodium vivax¹²12 Pincelli A, Cardoso MA, Malta MB, Johansen IC, Corder RM, Nicolete VC, et al.; MINA-Brazil Study Working Group. Low-level Plasmodium vivax exposure, maternal antibodies, and anemia in early childhood: population-based birth cohort study in Amazonian Brazil. PLoS Negl Trop Dis. 2021 Jul;15(7):e0009568. https://doi.org/10.1371/journal.pntd.0009568
https://doi.org/10.1371/journal.pntd.000... . The annual malaria incidence (API; number of laboratory-confirmed cases per 1,000 people per year)—one of the highest among Brazilian municipalities—was estimated at 231.9 in CZS in 2016¹²12 Pincelli A, Cardoso MA, Malta MB, Johansen IC, Corder RM, Nicolete VC, et al.; MINA-Brazil Study Working Group. Low-level Plasmodium vivax exposure, maternal antibodies, and anemia in early childhood: population-based birth cohort study in Amazonian Brazil. PLoS Negl Trop Dis. 2021 Jul;15(7):e0009568. https://doi.org/10.1371/journal.pntd.0009568
https://doi.org/10.1371/journal.pntd.000... . Mother-baby pairs were enrolled at pregnancy in public antenatal clinics, or at birth in the Women and Children's Hospital of Juruá Valley, the only maternity hospital of CZS. Informed consent forms were obtained at enrollment from mothers or caregivers (in the case of teenage mothers). The research protocols were approved by the Research Ethics Committee of the Faculdade de Saúde Pública da Universidade de São Paulo, Brazil (# 872.613, 2014; # 2.358.129, 2017).

During pregnancy, clinical and laboratory assessments were conducted by the research team in a subsample (n = 557) of the entire cohort (n =1,246). At delivery, the following data from both interviews and medical records were obtained: sociodemographic (maternal schooling, mothers’ self-reported skin color, whether the family was supported by the Bolsa Família Program, a conditional cash transfer program, and an assets-based wealth index used as a proxy of socioeconomic status) and maternal and perinatal data (such as parity, number of antenatal care visits, smoking and alcohol consumption during pregnancy, maternal hemoglobin concentration and body weight at delivery, gestational age and type of delivery, child's sex, and birthweight). From the prenatal card, information on maternal height (m) and pre-pregnancy weight (kg) were collected for all participants. Pre-pregnancy body mass index (BMI) was categorized as underweight (< 18.5 kg/m²), normal weight (18.5–24.9 kg/m²), overweight (25.0–29.9 kg/m²), or obese (≥ 30.0 kg/m²) as defined by the WHO. The difference between weight at delivery and pre-pregnancy weight was used to estimate maternal gestational weight gain (GWG). Based on pre-pregnancy BMI categories, GWG was classified as insufficient, adequate, or excessive following the Institute of Medicine 2009 guidelines¹³13 Institute of Medicine and National Research Council. Implementing guidelines on weight gain and pregnancy: Washington, DC: National Academy Press; 2013 [cited 2023 May 22]. Available from: https://www.nap.edu/catalog/18292/implementing-guidelines-on-weight-gain-and-pregnancy
https://www.nap.edu/catalog/18292/implem... .

After birth, clinical assessments with blood sample collection were performed at healthcare units when children were aged one, two, and five years, as described elsewhere¹⁰10 Cardoso MA, Matijasevich A, Malta MB, Lourenco BH, Gimeno SG, Ferreira MU, et al.; MINA-Brazil Study Group. Cohort profile: the maternal and child health and nutrition in Acre, Brazil, birth cohort study (MINA-Brazil). BMJ Open. 2020 Feb;10(2):e034513. https://doi.org/10.1136/bmjopen-2019-034513
https://doi.org/10.1136/bmjopen-2019-034... . Anthropometric measurements were performed in duplicate using standardized procedures. Child anthropometric indexes in z-scores were calculated according to age and sex following the WHO Child Growth Standards; stunting and overweight were defined as length or height for age in z-score < −2 and body mass index (BMI)-to-age > 2 z-scores, respectively¹⁴14 Onis M; WHO Multicentre Growth Reference Study Group. WHO Child Growth Standards based on length/height, weight and age. Acta Paediatr Suppl. 2006 Apr;450 S450:76-85. https://doi.org/10.1111/j.1651-2227.2006.tb02378.x
https://doi.org/10.1111/j.1651-2227.2006... .

At each childhood follow-up visit, structured questionnaires were administered to mothers or guardians to update data, including infant feeding practices, morbidities since birth, and other characteristics. Mothers reported whether the child was being breastfed (yes or no) and, if not, the age of weaning. Children who received breast milk with no other food or drink, except prescribed medicines, oral rehydration solutions, vitamins, and minerals, were considered exclusively breastfed. Continued breastfeeding (BF) was estimated in days and then classified as prolonged BF when ≥ 365 days. At the 1- and 2-year follow-up visits, information on complementary feeding was assessed by a structured food frequency questionnaire on the intake of foods and drinks in the previous day, as detailed elsewhere¹⁵15 Nogueira MB, Mazzucchetti L, Mosquera PS, Cardoso MA, Malta MB. Consumption of ultra-processed foods during the first year of life and associated factors in Cruzeiro do Sul, Brazil. Cien Saude Colet. 2022;27(2):725-36. https://doi:10.1590/1413-81232022272.47072020
https://doi:10.1590/1413-81232022272.470... . The following ultra-processed foods (UPF) were included in the questionnaire: industrialized yogurt, artificial fruit juice, soft drinks, candies, cookies, packaged savory snacks, hotdogs, and instant noodles, and “other UPF” (chocolate drinks, ice cream, jelly, cake, and industrialized soup). Then, the prevalence of UPF consumption was estimated based on the intake of at least one food from the category during the previous day.

Venous blood samples (10 mL for third-trimester pregnant women and 5 mL for children) were collected for measuring biochemical nutritional indicators during pregnancy and at the 1- and 2-year follow-up assessments; at the 5-year follow-up, only blood hemoglobin (Hb) concentration was measured. A Hb concentration < 110 g/L, measured in antenatal clinics using a portable Hemocue (Hb301; Angelholm, Sweden) hemoglobinometer defined anemia in pregnancy¹⁶16 World Health Organization. Haemoglobin concentrations for the diagnosis of anaemia and assessment of severity. Geneva: World Health Organization; 2011 [cited 2023 May 22]. Available from: http://www.who.int/vmnis/indicators/haemoglobin.pdf
http://www.who.int/vmnis/indicators/haem... . A Labtest SDH-20 cell counter (Labtest, Lagoa Santa, Brazil) was used to measure mothers’ Hb levels at the maternity ward. . An ABX Micro 60 cell counter (Horiba, Montpellier, France) was used to measure children's Hb concentrations at the 1- and 2-year follow-up visits; whereas at the 5-year follow-up, a portable Hemocue hemoglobinometer was used. Blood samples were protected from light and centrifuged within two hours after collection; serum and plasma samples were frozen at −20°C, shipped on dry ice to São Paulo, and kept at −70°C until further analyses. Plasma ferritin and soluble transferrin receptor concentrations were measured using enzyme immunoassay (Ramco, Houston, TX, USA). Liquid chromatography (HPLC) was used to measure serum concentrations of retinol as described elsewhere¹⁷17 Neves PA, Lourenço BH, Pincelli A, Malta MB, Souza RM, Ferreira MU, et al.; MINA-Brazil Study Group. High prevalence of gestational night blindness and maternal anemia in a population-based survey of Brazilian Amazonian postpartum women. PLoS One. 2019 Jul;14(7):e0219203. https://doi.org/10.1371/journal.pone.0219203
https://doi.org/10.1371/journal.pone.021... . Fluoroimmunoassay (PerkinElmer; Wallac Oy, Turku, Finland) was used to measure serum folate concentrations. An IMMAGE Immunochemistry System (Beckman Coulter, Brea, CA, USA) was used to determine C-reactive protein. Analyses were subject to internal and external quality control with routine use of blind samples for each run, with coefficients of variation < 7%.

Cut-off values established by the WHO were adopted to define vitamin deficiencies: serum retinol concentrations < 0.7 μmol/L for vitamin A deficiency (VAD) and < 1.05 μmol/L for vitamin A insufficiency (VAI)¹⁸18 World Health Organization. Global prevalence of vitamin A defciency in populations at risk: Geneva: World Health Organization; 2009 [cited 2023 May 22]. Available from: https://apps.who.int/iris/bitstream/handle/10665/44110/9789241598019_eng.pdf
https://apps.who.int/iris/bitstream/hand... , and serum folate concentration < 3 ng/mL (pregnant women) or < 4 ng/mL (children) for folate deficiency¹⁹19 World Health Organization. Serum and red blood cell folate concentrations for assessing folate status in populations. Geneva: World Health Organization; 2012 [cited 2023 May 22]. Available from: http://apps.who.int/iris/bitstream/handle/10665/75584/WHO_NMH_NHD_EPG_12.1_eng.pdf;jsessionid=EE15CBE23687EB40C896BB4D3FF41B28?sequence=1
http://apps.who.int/iris/bitstream/handl... . Iron deficiency (ID) was defined as plasma ferritin concentration < 12 μg/L (children) or < 15 μg/L (pregnant women), combined with a soluble transferrin receptor concentration > 8.3 mg/L, or plasma ferritin < 30 μg/L combined with a C-reactive protein concentration ≥ 5 mg/L (suggestive of acute inflammation)²⁰20 World Health Organization. WHO guideline on use of ferritin concentrations to assess iron status in individuals and populations. Geneva: World Health Organization; 2020 [cited 2023 May 22]. Available from: https://www.who.int/publications/i/item/9789240000124
https://www.who.int/publications/i/item/... .

Malaria during pregnancy was diagnosed by microscopy during visits to health units and/or by real-time polymerase chain reaction (PCR) at delivery²¹21 Baroni L, Pedroso M, Barcellos C, Salles R, Salles S, Paixão B, et al. An integrated dataset of malaria notifications in the Legal Amazon. BMC Res Notes. 2020 Jun;13(1):274. https://doi.org/10.1186/s13104-020-05109-y
https://doi.org/10.1186/s13104-020-05109... . Malaria in young children was diagnosed by microscopy during visits to health units. Case notifications during pregnancy and the first two years of life were retrieved from the electronic database of the Brazilian Ministry of Health (SIVEP Malaria) using a previously described linkage strategy¹²12 Pincelli A, Cardoso MA, Malta MB, Johansen IC, Corder RM, Nicolete VC, et al.; MINA-Brazil Study Working Group. Low-level Plasmodium vivax exposure, maternal antibodies, and anemia in early childhood: population-based birth cohort study in Amazonian Brazil. PLoS Negl Trop Dis. 2021 Jul;15(7):e0009568. https://doi.org/10.1371/journal.pntd.0009568
https://doi.org/10.1371/journal.pntd.000... .

Anemia, nutritional deficiencies, and malaria diagnosed during the follow-up visits were treated by research physicians. Children with anemia were prescribed ferrous sulfate (dose: 3–6 mg of elemental iron/kg per day up to 60 mg/day for four months) for presumptive treatment of iron deficiency, but adherence to this treatment was not evaluated.

Main Outcome Measures

Childhood anemia was defined as Hb < 110 g/L for children < 5 years. At the 5-year follow-up visit, Hb < 115 g/L defined anemia for children ≥ 5 years. Severe anemia was classified as Hb < 70 g/L and < 80 g/L for children < 5 and ≥ 5 years, respectively¹⁶16 World Health Organization. Haemoglobin concentrations for the diagnosis of anaemia and assessment of severity. Geneva: World Health Organization; 2011 [cited 2023 May 22]. Available from: http://www.who.int/vmnis/indicators/haemoglobin.pdf
http://www.who.int/vmnis/indicators/haem... .

Data Analysis

Maternal and child characteristics were described using absolute frequencies and proportions (%) with 95% confidence intervals (CI). Mean and SD for age and median values and interquartile ranges (IQR, 25^th and 75^th percentiles) for biochemical nutritional indicators were calculated. Pearson χ² or Fisher exact tests were used to compare proportions.

Relative risks (RR) with 95%CI were estimated using multiple Poisson regression models with robust variance to identify factors associated with childhood anemia at each follow-up visit and with “persistent anemia” (children who were anemic in both the 1-year and the 2-year visits). Multiple adjusted linear regression coefficients (β) were also calculated to describe predictors for concentrations of Hb (dependent variable) at each follow-up assessment. Variables associated with the outcomes at a significance level < 20% in unadjusted analysis were entered in multiple regression models. A hierarchical approach was used based on conceptual frameworks⁷7 Victora CG, Christian P, Vidaletti LP, Gatica-Domínguez G, Menon P, Black RE. Revisiting maternal and child undernutrition in low-income and middle-income countries: variable progress towards an unfinished agenda. Lancet. 2021 Apr;397(10282):1388-99. https://doi.org/10.1016/S0140-6736(21)00394-9
https://doi.org/10.1016/S0140-6736(21)00... ,²²22 Cardoso MA, Scopel KK, Muniz PT, Villamor E, Ferreira MU. Underlying factors associated with anemia in Amazonian children: a population-based, cross-sectional study. PLoS One. 2012;7(5):e36341. https://doi.org/10.1371/journal.pone.0036341
https://doi.org/10.1371/journal.pone.003... to select distal (demographic and socio-economic factors), underlying (antenatal care, obstetric and birth characteristics), and immediate (child feeding practices, nutritional status, and morbidities) determinants of childhood anemia in the final adjusted models (Figure 1). At each level of determination, covariates were retained in the model if they were associated with the outcome at P < 0.10 and/or for ordinal variables with more than two categories if they followed a dose-response pattern. Missing values in categorical covariates were maintained in the model by creating a new missing-value category. All P-values reported are two-tailed and the significant level was set at P < 0.05. Statistical analyses were performed using Stata version 15.0 (StataCorp, College Station, TX, USA).

RESULTS

At baseline, we obtained data from 1,246 participants of the MINA Brazil cohort. After exclusion of 22 twins, a total of 1,224 mother-child pairs were eligible for the present analysis. Of these, 79 missed all post-natal follow-up visits and six children died up to five years of age (Figure 2). Children participating from birth to different follow-up assessments and those lost to follow-up over time until the 5-year assessment (n = 514) had similar perinatal characteristics regarding sex, gestational age, preterm birth, and birth weight, but differed significantly in the proportion of children from poorest families (36.2% versus 44.1%) and of mothers with ≤ 9 years of schooling (26.4% versus 47.5%), respectively, with P < 0.01 (c² test) for both.

The overall prevalence of childhood anemia declined from 42% to 13% and 5% at 1-, 2-, and 5-year assessments, respectively. A total of two cases (0.3%) of severe anemia (Hb < 70 g/L) were observed at the 1-year follow-up; the prevalence of moderate anemia was 14.2%, 1.5%, and 1.5% at 1, 2, and 5 years of age, respectively (Table 1). In crude analysis, the lowest wealth index, lowest maternal age, lowest schooling, prolonged breastfeeding, and consumption of UPF were significantly associated with anemia at one year of age. At two years, children in the lowest wealth index, born to a multiparous mother, with < 6 antenatal care visits, and history of one or more malaria infections since birth were at higher risk for anemia. At five years, only prolonged breastfeeding was significantly associated with lower risk of anemia.

Table 2 shows the mean values of age (SD), frequencies of the use of iron and/or vitamin supplements, prevalence (95%CI) of nutritional deficiencies, and interquartile ranges (IQR) of biochemical parameters at different waves of the entire cohort. Among pregnant and parturient women, the most frequently used supplements were iron (30–60 mg/day) and folic acid (5 mg/day), following national antenatal care guidelines. At delivery, similar frequencies (around 30%) of underweight and excessive GWG were observed. Anemia was common among pregnant women (17%) and parturients (38%). Inadequacies of iron or vitamin A status were frequent among pregnant women and children under two years; folic acid deficiency was uncommon, with prevalence < 1% in mothers and children. Combined iron and vitamin A deficiencies were more common at two years (6.7%).

ID (adjusted RR [RR_a] = 2.19; 95%CI: 1.84–2.60) and consumption of UPF (RR_a = 1.56; 95%CI: 1.14–2.13) were associated with elevated risk of anemia at one year, after adjustment for maternal schooling in multiple regression analysis (Table 3). At the 2-year follow-up visit, maternal anemia at delivery (RR_a = 1.67; 95%CI: 1.17–2.39), malaria in the first 2 years of life (RR_a = 2.25; 95%CI: 1.30–3.87), and ID (RR_a = 2.15; 95%CI: 1.47–3.15) were significantly associated with increased risk of anemia, after adjustment for child's age and household wealth index. Risk of persistent anemia in the 1- and 2-year follow-up visits (n = 53) was positively associated with male sex (RR_a = 1.87; 95%CI: 1.05–3.31), malaria during the first two years of life (RR_a = 4.30; 95%CI: 2.33–7.94), prolonged breastfeeding (RR_a = 2.48; 95%CI: 1.11–5.51), and vitamin A insufficiency (RR_a = 2.03; 95%CI: 1.16–3.54); consumption of meat emerged as a protective factor (RR_a = 0.52; 95%CI: 0.29– 0.95). At 5 years, only maternal anemia at delivery was positively associated with risk of anemia (RR_a = 2.32; 95%CI: 1.11– 4.84), whereas prolonged breastfeeding was inversely associated with risk of anemia (RR_a = 0.45; 95%CI: 0.22– 0.94).

Predictors of Hb concentrations during childhood were also explored (Table 3). At one year, maternal schooling was positively associated with Hb concentration, whereas ID and VAI were negatively associated. At two years, children of families in the highest wealth stratum, born to primiparous mothers, and those who consumed beans had higher Hb concentrations, whereas maternal anemia at delivery, malaria in the first 2 years, and iron deficiency were negatively associated with Hb concentration. At five years, prolonged breastfeeding was positively associated with Hb concentration; conversely, maternal anemia, consumption of UPF at one year, and malaria in the first two years of life showed a negative association.

DISCUSSION

In this study, we addressed some of the most likely causes of childhood anemia in the Amazon. We showed that iron deficiency (ID) is still very common, affecting 43% of local pregnant women and 38% of children at one year of age, but its contribution to anemia seems to vary with age. Accordingly, we diagnosed ID in 51% of pregnant women and 62% of the children aged one who were found to be anemic, which was observed in only 32% of the anemic children aged two. VAD and VAI were also common during pregnancy (6% and 20%, respectively) and in children aged two (25% and 42%, respectively), which was substantially less frequent in children aged one (2% and 10%, respectively), when childhood anemia is most prevalent. Evidence that VAI or VAD contributes to anemia in our population is twofold: we found a significant association between VAI and elevated risk of persistent anemia in early childhood, as well as a negative impact of VAI on Hb levels at the age of one year.

Children born to mothers who were anemic at delivery were more likely to be anemic at the ages of two and five after adjusting for potential confounders. Although this association is likely to be causal, residual confounding must also be considered since mother-child pairs sharing the same household are similarly exposed to poverty, food insecurity, and other key contributors to nutritional deficiencies and anemia. We highlight that recent evidence shows that anemia in pregnancy increases the risk of ID and anemia in infants, suggesting that the benefits of preventing anemia in pregnancy might extend to the offspring by reducing anemia risk in early childhood²³23 Davidson EM, Simpson JA, Fowkes FJ. The interplay between maternal-infant anemia and iron deficiency. Nutr Rev. 2023 Mar;81(4):480-91. https://doi.org/10.1093/nutrit/nuac066
https://doi.org/10.1093/nutrit/nuac066... . Our results reinforce the argument for routine iron supplementation in pregnancy as advocated by WHO in areas with a prevalence > 40%¹⁶16 World Health Organization. Haemoglobin concentrations for the diagnosis of anaemia and assessment of severity. Geneva: World Health Organization; 2011 [cited 2023 May 22]. Available from: http://www.who.int/vmnis/indicators/haemoglobin.pdf
http://www.who.int/vmnis/indicators/haem... .

Folate deficiency was rare among MINA-Brazil cohort participants, most likely due to folic acid fortification of wheat flour implemented in Brazil in 2004. Brazil has adopted the fortification of wheat and maize flours with 4.2 mg of iron and 150 mg of folic acid added per 100 g. However, the effectiveness of iron fortification of flour in preventing anemia seems to be poor among young children²⁴24 Assunção MC, Santos IS, Barros AJ, Gigante DP, Victora CG. Effect of iron fortification of flour on anemia in preschool children in Pelotas, Brazil]. Rev Saude Publica. 2007 Aug;41(4):539-48. https://doi.org/10.1590/S0034-89102006005000031
https://doi.org/10.1590/S0034-8910200600... . The transition to complementary foods as children begin to consume household diets is associated with insufficient iron intake, combined with low dietary intake of other micronutrients⁷7 Victora CG, Christian P, Vidaletti LP, Gatica-Domínguez G, Menon P, Black RE. Revisiting maternal and child undernutrition in low-income and middle-income countries: variable progress towards an unfinished agenda. Lancet. 2021 Apr;397(10282):1388-99. https://doi.org/10.1016/S0140-6736(21)00394-9
https://doi.org/10.1016/S0140-6736(21)00... . Thus, since 2011, the use of multiple micronutrients in powder (MNP) has been recommended by WHO as a home-based strategy to prevent and control childhood anemia. In 2014, the Brazilian Ministry of Health launched the Multiple Micronutrient Powders For Fortification of Foods Consumed by Infants and Children (NUTRISUS)) based on the results of a multicenter pragmatic trial following WHO guidelines in settings where the prevalence of anemia in young children is 20% or higher²⁵25 Cardoso MA, Augusto RA, Bortolini GA, Oliveira CS, Tietzman DC, Sequeira LA, et al. Effect of providing multiple micronutrients in powder through primary healthcare on anemia in young brazilian children: a multicentre pragmatic controlled trial. PLoS One. 2016 Mar;11(3):e0151097. https://doi.org/10.1371/journal.pone.0151097
https://doi.org/10.1371/journal.pone.015... . However, since 2016, political and economic crises in Brazil have drastically reduced access to many primary health programs, which could have contributed to the high prevalence of anemia in young children reported in this study.

Other important actions to prevent and control anemia in young children in Brazil include the promotion of exclusive breastfeeding during the first six months of life and of a healthy and timely introduction of complementary feeding, which should be encouraged in primary healthcare settings. Among children with persistent anemia from one to two years, our results reinforce the importance of promoting continued breastfeeding with nutritious complementary diets and adequate treatment of childhood infections such as malaria. In a previous analysis of the MINA-Brazil cohort, children exposed to gestational malaria and breastfed for at least 12 months had a decreased risk for malaria infection during the first two years of life²⁶26 Pincelli A, Cardoso MA, Malta MB, Nicolete VC, Soares IS, Castro MC, et al.; MINA-Brazil Study Working Group. Prolonged breastfeeding and the risk of plasmodium vivax infection and clinical malaria in early childhood: a birth cohort study. Pediatr Infect Dis J. 2022 Oct;41(10):793-9. https://doi.org/10.1097/INF.0000000000003618
https://doi.org/10.1097/INF.000000000000... . Breastfeeding protects young children from infections with the passive transfer of immunoglobulins and other bioactive substances²⁶26 Pincelli A, Cardoso MA, Malta MB, Nicolete VC, Soares IS, Castro MC, et al.; MINA-Brazil Study Working Group. Prolonged breastfeeding and the risk of plasmodium vivax infection and clinical malaria in early childhood: a birth cohort study. Pediatr Infect Dis J. 2022 Oct;41(10):793-9. https://doi.org/10.1097/INF.0000000000003618
https://doi.org/10.1097/INF.000000000000... , which could be one possible explanation for the association of prolonged breastfeeding and decreased risk for anemia at age five observed in this study.

To the best of our knowledge, this is the first study to report the significant association between UPF consumption with anemia at age one. A recent systematic review described a higher participation of UPF in children's diet associated with other maternal-child outcomes, such as an increase in weight gain, adiposity measures, overweight, early weaning, lower diet quality, metabolic alterations, diseases, and ingestion of plastic from packaging²⁷27 Oliveira PG, Sousa JM, Assunção DG, Araujo EK, Bezerra DS, Dametto JF, et al. Impacts of consumption of ultra-processed foods on the maternal-child health: a systematic review. Front Nutr. 2022 May;9:821657. https://doi.org/10.3389/fnut.2022.821657
https://doi.org/10.3389/fnut.2022.821657... . As defined by the NOVA classification, UPF are industrial formulations of substances derived from foods with little or no whole food with added colorings, flavorings, emulsifiers, thickeners, and other cosmetic additives to make them palatable or even hyperpalatable²⁸28 Monteiro CA, Cannon G, Lawrence M, Costa Louzada MD, Pereira Machado P. Ultra-processed foods, diet quality, and health using the NOVA Classification System. Rome: FAO; 2019 [cited 2023 May 22]. Available from: https://www.fao.org/3/ca5644en/ca5644en.pdf
https://www.fao.org/3/ca5644en/ca5644en.... . In a previous study, the introduction of UPF in complementary feeding was inversely associated with the duration of continued breastfeeding²⁹29 Soares MM, Juvanhol LL, Ribeiro SA, Franceschini SD, Araújo RM. Prevalence of processed and ultra-processed food intake in Brazilian children (6-24 months) is associated with maternal consumption and breastfeeding practices. Int J Food Sci Nutr. 2021 Nov;72(7):978-88. https://doi.org/10.1080/09637486.2021.1885016
https://doi.org/10.1080/09637486.2021.18... , providing additional evidence to avoid UPF consumption in childhood.

This study presents some limitations. First, MINA-Brazil cohort participants were not screened for helminthic infections, inherited hemolytic disorders, and other non-nutritional conditions that may be associated with anemia in tropical environments. Accordingly, 4–10% of Amazonian children under the age five tested in population-based cross-sectional surveys harbor one or more soil-transmitted intestinal helminths that may cause or aggravate anemia²²22 Cardoso MA, Scopel KK, Muniz PT, Villamor E, Ferreira MU. Underlying factors associated with anemia in Amazonian children: a population-based, cross-sectional study. PLoS One. 2012;7(5):e36341. https://doi.org/10.1371/journal.pone.0036341
https://doi.org/10.1371/journal.pone.003... . However, their contribution to anemia among Amazonian children, although not negligible, seems to be much less pronounced than that in African or Asian populations²²22 Cardoso MA, Scopel KK, Muniz PT, Villamor E, Ferreira MU. Underlying factors associated with anemia in Amazonian children: a population-based, cross-sectional study. PLoS One. 2012;7(5):e36341. https://doi.org/10.1371/journal.pone.0036341
https://doi.org/10.1371/journal.pone.003... . Glucose-6-phosphate dehydrogenase (G6PD) deficiency, an X-linked recessive disorder, is the most common inherited cause of hemolytic anemia in the Amazon. It affects 8.3% of the male population of Acre State³⁰30 Nascimento JR, Brito-Sousa JD, Almeida AC, Melo MM, Costa MR, Barbosa LR, et al. Prevalence of glucose 6-phosphate dehydrogenase deficiency in highly malaria-endemic municipalities in the Brazilian Amazon: a region-wide screening study. Lancet Reg Health Am. 2022 May;12:100273. https://doi.org/10.1016/j.lana.2022.100273
https://doi.org/10.1016/j.lana.2022.1002... , who may develop mild to severe hemolysis when exposed to dietary triggers and some medicines, including the locally used antimalarial primaquine. Nevertheless, G6PD deficiency has not emerged as a significant predictor of Hb levels or risk of anemia in Amazonian children²²22 Cardoso MA, Scopel KK, Muniz PT, Villamor E, Ferreira MU. Underlying factors associated with anemia in Amazonian children: a population-based, cross-sectional study. PLoS One. 2012;7(5):e36341. https://doi.org/10.1371/journal.pone.0036341
https://doi.org/10.1371/journal.pone.003... . Second, the prevalence of iron deficiency and anemia may have been underestimated in MINA-Brazil cohort participants over time because children lost to follow-up were more likely to live in the poorest households, being more exposed to food insecurity, malaria, and other contributors to nutritional deficiencies and anemia. A range of other conditions (e.g., tropical infections, chronic inflammation, and inherited hemolytic disorders), some of them not addressed by this study, may have contributed to the high prevalence of anemia in Amazonian populations.

CONCLUSION

Although our study population has access to antenatal care, prompt malaria diagnosis and treatment, and iron and vitamin A supplementation, anemia in MINA-Brazil cohort participants remains common during pregnancy and early childhood, especially at the age one. Moreover, ID and consumption of UPF during age one were associated with elevated risk of anemia. As a multifactorial public health problem, different possible causes should be considered in all efforts and programs to achieve the SDGs targets in this region.

Acknowledgments:

We would like to thank all women and children who have taken part in the MINA-Brazil Study and healthcare professionals at the Maternity Hospital, Municipal Health Secretariat, and primary healthcare units of Cruzeiro do Sul. Members of the MINA-Brazil Working Group: Marly Augusto Cardoso (PI), Alicia Matijasevich, Bárbara Hatzlhoffer Lourenço, Jenny Abanto, Maíra Barreto Malta, Marcelo Urbano Ferreira, Paulo Augusto Ribeiro Neves (University of São Paulo, São Paulo, Brazil); Ana Alice Damasceno, Bruno Pereira da Silva, Rodrigo Medeiros de Souza (Federal University of Acre, Cruzeiro do Sul, Brazil); Simone Ladeia-Andrade (Oswaldo Cruz Institute, Fiocruz, Rio de Janeiro, Brazil), Marcia Caldas de Castro (Harvard T.H. Chan School of Public Health, Boston, USA).

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