Jiang JF, Jiang JB, Zhu HS, et al. Combined treatment with vitamin A and iron to prevent piglet anemia. J Swine Health Prod. 2009;17(1):22–27

Cite as: Jiang JF, Jiang JB, Zhu HS, et al. Combined treatment with vitamin A and iron to prevent piglet anemia. J Swine Health Prod. 2009;17(1):22–27.
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Keywords: swine, piglet anemia, vitamin A, iron
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Received: October 15, 2007
Accepted: June 15, 2008

Anemia is the most prevalent nutritional deficiency in the world.¹ Both iron and vitamin A deficiencies were independent risk factors for anemia among Marshall Islands preschool children.² Attempts to improve iron status have been thwarted by deficiency of and adverse interaction with other micronutrients.¹ Confinement-reared pigs develop iron deficiency anemia (hypochromic, microcytic anemia) early in life. Anemia occurs because piglets are born with unusually small iron stores, milk contains low levels of iron, and pigs have a very rapid growth rate.³ Anemia interferes with growth, and affected pigs are listless and more susceptible to infectious diseases than are normal pigs.

Supplemental vitamin A increased hemoglobin levels and packed cell volume (PCV) in pregnant women with deficient iron status or marginally deficient or deficient vitamin A status.⁴ Vitamin A supplementation thereby contributed to control of nutritional anemia, and there was a synergistic interaction between vitamin A and iron in combined therapy. Vitamin A supplementation of pregnant women was associated with higher birth weight and lower prevalence of anemia among their infants.⁵ Vitamin A deficiency appears to be related to the pathogenesis of anemia by several biological mechanisms, eg, improvement of growth and differentiation of erythrocyte progenitor cells, promotion of immunity to infection, less severe anemia of infection, and mobilization of iron stores from tissues.^6,7

Concomitant supplemental vitamin A and ferrous sulfate promoted the hematopoietic effect of iron supplementation in young male rats.⁸ Concomitant supplementation of vitamin A enhanced the response to weekly supplementation of iron and folic acid in anemic teenagers in urban Bangladesh.⁹ A relationship between vitamin A and iron may have relevance in the treatment of nutritional anemia, and some studies suggest that supplementation with both vitamin A and iron is superior to iron alone in treating nutritional anemia in humans and rats.¹⁰ The aim of this study was to evaluate combined treatment with iron and vitamin A in preventing piglet anemia.

Materials and methods

Study animals

Ninety-six piglets (Duroc × Large White × Landrace), born to eight crossbred multiparous sows (third or fourth parturition; Large White × Landrace) inseminated with semen from the same boar, were used in this study. All experimental procedures, care, and handling of animals were conducted according to guidelines on the care and use of animals for scientific purposes.¹¹ All animal experiments were approved by the Local Ethics Committee of Animal Research (permit no. C 16/6).

Experimental design

Piglets were assigned to eight blocks (12 piglets per block) on Day 0 (the day of birth) according to the following criteria: litter, gender, and weight. Each animal in a block was randomly allotted to one of 12 groups which comprised three treatments (four replicates per treatment). Treatments consisted of the control (no supplemental iron; n = 32); intramuscular (IM) injection of 200 mg iron as iron dextran (Bestar Laboratories Ltd, Shanghai, China) on Day 2 (n = 32); and IM injection of 200 mg iron as iron dextran plus oral administration of 2000 IU vitamin A as vitamin A palmitate (Beijing Zhongnongjianuo Technology Co Ltd, Beijing, China) on Day 2 (n = 32).

Immediately after birth, all piglets were encouraged to suckle to ensure ingestion of colostrum, then litter size was equalized to 12 by cross-fostering. Within 12 hours after birth, piglets in each litter were stratified according to weight and randomly assigned by weight to treatment groups. Piglets within each litter were equally allotted to the 12 groups; thus, each sow nursed piglets belonging to each of the 12 groups. Piglets were individually identified with ear tags.

Twelve pigs (four randomly selected pigs from each treatment) were euthanized by electrocution on Day 21. Applying the electrodes to the head of pigs for a minimum of 5 seconds resulted in instantaneous loss of consciousness (collapse, immediate mydriasis, no vocalization), and subsequently applying the electrodes to the thorax of pigs for a minimum of 15 seconds caused cardiac arrest within 1 minute. Death was confirmed and electrodes were removed from the thorax. The liver and spleen were removed as soon as possible after euthanasia and stored at -20°C until iron analysis 10 days later.

Housing and feeding

The farrowing barn housed piglets and their sows in farrowing crates with thermostatically controlled creep boxes and plastic-coated slatted floors. Air temperature of the farrowing pens was maintained at approximately 18°C through the experiment. Temperature in the creep boxes was maintained at approximately 32°C using a 250-watt electric heat lamp from Day 0 to Day 6. Temperature in the creep area was reduced to 30.5°C, 29°C, 27°C, and 25°C on Days 7, 11, 15, and 19 by changing to bulbs of 240, 225, 210, and 195 watts, respectively. On Day 1, all piglets were processed according to standard commercial practices, including teeth clipping, tail docking, and ear notching. During the previous gestation and lactation, the dams of the experimental pigs were fed a corn-soybean meal-based diet formulated to meet National Research Council¹² nutrient requirement estimates (Table 1). During the study, sows and piglets had ad libitum access to feed and water.

Parameters measured

Piglets were individually weighed on an electronic scale accurate to 0.1 kg within 12 hours after birth and on Day 21. Blood samples collected in EDTA tubes were obtained via vena cava puncture on Days 1, 7, 14, and 21. One portion of the blood from each pig was immediately prepared for hemoglobin and hematocrit determination. The second EDTA tube of blood from each pig was chilled to 4°C and centrifuged at 2000g. Plasma was harvested and stored at -20°C until iron and total iron-binding capacity were determined 2 days later.

Iron was measured in plasma, liver, and spleen using atomic absorption spectrophotometry. Liver and spleen samples were prepared for iron analysis by blotting dry, drying in a forced-air oven at 100°C, then wet-ashing with nitric acid in a microwave oven as described by Kegley et al.³ Hemoglobin and total iron-binding capacity were measured by using commercially available test kits (Sigma Chemical Co, St Louis, Missouri). Hematocrit was measured using microcapillary tubes as described by Kegley et al.³

Deaths were recorded in each group. Mortality for each group was calculated as the total number of pigs at the beginning of the experiment minus the number of dead piglets at the end of the experiment, expressed as a percentage of the total number of pigs.

Data analysis

Data were analyzed by analysis of variance (ANOVA) using PROC MIXED of SAS (SAS 9.0, SAS Institute Inc, Cary, North Carolina). Variables included weight gain, mortality, iron concentrations in the liver and spleen, hematocrit, hemoglobin concentration, plasma iron concentration, and total iron-binding capacity. The model for weight gain, mortality, and iron concentrations in the liver and spleen included treatment and litter as fixed effects. The model for hematocrit, hemoglobin concentration, plasma iron concentration, and total iron-binding capacity included treatment, litter, day, and the interaction of treatment and day as fixed effects. When treatment effect was a significant source of variation, differences were determined using the DIFF option of SAS. Least squares means were calculated for each independent variable. Statistical significance was set at P < .05 for all statistical tests.

Results

Weight gains during the study were greater (P < .05) for pigs treated either with iron alone or with iron and vitamin A than for the controls (Table 2). Weight gains did not differ significantly between pigs treated with iron alone and pigs treated with both iron and vitamin A (P > .05).

Mortality during the study was higher in the control group (P < .05) than in the treated groups (Table 2). Mortality did not differ significantly between pigs treated with iron alone and pigs treated with both iron and vitamin A (P > .05).

On Day 21, iron concentrations in liver and spleen samples from pigs that received no iron were lower (P < .05) than concentrations in pigs treated with iron (Table 2). Concentration of iron in the liver was 10% higher in pigs treated with both iron and vitamin A than in pigs treated with iron alone (P < .05; Table 2). Concentration of iron in the spleen did not differ significantly between pigs that were treated with iron alone and pigs treated with both iron and vitamin A (P > .05; Table 2).

Hematocrits did not differ significantly on Day 1 (Table 3). On Days 7, 14, and 21, hematocrits were lower in control pigs than in either group of pigs treated with iron (P < .05). On Day 7, hematocrits were 10.3% higher (P < .05) in pigs treated with both iron and vitamin A than in pigs treated with iron alone. On Days 14 and 21, hematocrits did not differ significantly between pigs treated with iron alone and pigs treated with both iron and vitamin A (P > .05).

Hemoglobin concentrations did not differ significantly on Day 1 (Table 4). On Days 7, 14, and 21, hemoglobin concentrations were lower in control pigs (P < .05) than in either group of pigs treated with iron. On Days 7, 14, and 21, hemoglobin concentrations were 11.2%, 10.6%, and 10.7% higher, respectively, in pigs treated with both iron and vitamin A than in pigs treated with iron alone (P < .05).

Plasma iron concentrations did not differ significantly on Day 1 (Table 5). Plasma iron concentrations were lower in controls than in pigs treated with iron on Days 7, 14 and 21 (P < .05). On Days 7 and 14, plasma iron concentrations in pigs treated with both iron and vitamin A were higher (P < .05) than those in pigs treated with iron alone. On Day 21, plasma iron concentrations did not differ significantly between pigs treated with iron alone and pigs treated with both iron and vitamin A (P > .05).

Total iron-binding capacity was higher (P < .05) in control pigs than in pigs treated with iron on Days 7, 14, and 21 (Table 6). On Day 7, total iron-binding capacity did not differ significantly between pigs treated with iron alone and pigs treated with both iron and vitamin A. On Days 14 and 21, total iron-binding capacity was lower in pigs treated with both iron and vitamin A than in pigs treated with iron alone (P < .05).

The effect of day on hematocrit, hemoglobin concentration, plasma iron concentration, and total iron-binding capacity was significant (P < .05). The treatment × day interaction was significant for hematocrit and plasma iron concentration (P < .05), but not for hemoglobin concentration and total iron-binding capacity (P > .05).

Discussion

Iron deficiency is the main cause of piglet anemia.¹³ Many researchers have proved that newborn pigs need supplemental iron, and administration of iron to the neonatal pig has been a standard practice in many parts of the world for many years. In the present study, weight gains between Day 0 and Day 21 were higher in pigs that were treated with iron on Day 2 than in pigs not treated with iron. Measures of iron nutrition, including hemoglobin concentration, hematocrit, plasma iron concentration, total iron-binding capacity, and concentrations of iron in liver and spleen were all significantly better in pigs that were treated with iron than in pigs not treated with iron, in agreement with other studies.^3,14,15

Intramuscular injection of iron in newborn pigs is effective in preventing piglet anemia in practice. In this study, intramuscular injection of iron combined with oral vitamin A was more effective than treatment with iron alone. Hemoglobin and hematocrits levels are sensitive criteria for evaluating body biological response to iron. Plasma iron concentration is the indicator of iron deficiency, and total iron-binding capacity is the important index in iron metabolism, representing the ability of transferrin to carry iron in the blood.¹⁶ In this study, hemoglobin concentration, hematocrit, plasma iron concentration, and iron concentrations in liver and spleen were higher and total iron-binding capacity was lower in pigs treated with both iron and vitamin A than in pigs treated with iron alone, showing that body iron status was better when piglets were treated with both iron and vitamin A. These data suggest that a combination of iron and vitamin A is more effective than iron alone in preventing piglet anemia. To the authors’ knowledge, there are no previously published reports concerning the combination of iron and vitamin A in preventing anemia in pigs.

In a study in humans with low vitamin A status or deficient iron status, supplemental vitamin A was associated with higher hemoglobin level and hematocrit, thereby contributing to control of nutritional anemia, and there was a synergistic interaction between vitamin A and iron in combined therapy.⁴ Iron in combination with vitamin A was more effective than iron alone in treating low iron status in rats that had lower blood hemoglobin concentration, hematocrit, and erythrocyte count after being fed a diet deficient in both iron and retinol.⁸ Mwanri et al¹⁷ conducted a randomized controlled trial to study the effects of dietary supplements on anemic children, using vitamin A alone, iron and vitamin A, iron alone, or placebo, administered in a double-blind design for 3 months. All supplements were administered in corn-based gruel. Results showed that after 3 months, mean hemoglobin concentration was higher by 13.5 g per L in children receiving vitamin A alone, compared with 3.5 g per L in the placebo treatment group (P < .0001). In addition, after 3 months, the mean body weight of children receiving vitamin A alone was higher by 0.6 kg, compared with 0.2 kg for the placebo treatment (P < .0001), and mean height for children receiving vitamin A alone was higher by 0.4 cm compared with 0.1 cm for the placebo treatment (P = .0009). However, in the group of children who received both vitamin A and iron supplementation, mean change from baseline was better in all indicators compared with the placebo treatment (mean change in hemoglobin 18.5 g per L, P < .0001; mean change in body weight 0.7 kg, P < .0001; and mean change in height 0.4 cm, P < .0001). The authors concluded that, in developing countries, vitamin A supplementation may have a useful role in combating anemia, as well as in improving children’s growth.

Zhang et al¹⁸ found that in broiler chickens, iron concentration in liver decreased and iron concentration in serum increased with an increase in dietary supplemental vitamin A. Duodenum iron concentration, tibia iron concentration, and erythrocyte count increased significantly with higher dietary supplemental vitamin A (P < .01), indicating that vitamin A can enhance iron metabolism.

The mechanism by which vitamin A alleviates iron deficiency anemia is not clear. There are several hypotheses. Vitamin A may form a complex with iron, maintaining its solubility in the intestinal lumen and preventing the inhibitory effects of phytates and polyphenols on iron absorption,^19,20 but Sajedianfard et al²¹ concluded that the therapeutic effect of vitamin A in iron-deficiency anemia is probably not associated with its influence on iron absorption from the gastrointestinal tract. The study of Amine et al²² showed that vitamin A deficiency impairs erythropoiesis, but Roodenburg et al²³ found no evidence that vitamin A deficiency affected erythropoiesis and erythrocyte turnover. Transcription of the transferrin gene in vitro is stimulated by vitamin A,^24,25 suggesting that vitamin A is involved in synthesis of the glycosyl moieties of the transferrin molecule.

Vitamin A seems to be related to the pathogenesis of anemia by various biological mechanisms, such as enhancing the growth and differentiation of erythrocyte progenitor cells, potentiating immunity to infection and reducing the anemia of infection, and mobilizing iron stores from tissues.⁷

There was a significant positive correlation between plasma retinol and plasma iron in pregnant women.²⁶ By measuring plasma retinol concentrations of piglets before and after suckling, Hakansson et al²⁷ established that piglets were born with low levels of retinol (0.07 mg per L), that transfer of vitamin A via the placenta appeared to be limited, and that colostrum was a major means of retinol transfer to the young piglet. Davila et al²⁸ reported that, in rats, vitamin A concentration in colostrum on day 1 of lactation did not vary with maternal vitamin A intake during pregnancy; however, the concentration of vitamin A in milk increased with increasing maternal vitamin A intake during lactation. In swine, the highest mean concentrations of retinol were found in colostrum, while retinol concentration in sow milk decreased by 71% during the first week postpartum and remained relatively stable thereafter.²⁷ These results^17,27,28 indicate that transfer of vitamin A via the placenta is limited, that piglets are born with small vitamin A stores, that colostrum is the main source of vitamin A for newborn piglets, that colostrum vitamin A concentration is not affected by maternal vitamin A intake during pregnancy, and that the vitamin A status of piglets is influenced by maternal vitamin A intake during lactation.

Implications

• Piglets have better iron nutrition status when provided with both iron and vitamin A than when treated with iron alone.
• Vitamin A may promote the role of iron in preventing piglet anemia.
• Under the conditions of this study, a combination of vitamin A and iron is more effective than iron alone in preventing piglet anemia.

Acknowledgments

The authors thank Feng Wang, Yinghua Shi, and Xianghua Yan for their assistance. The China National Research Foundation is acknowledged for financial support.

References

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