Effects of the in ovo injection of vitamin D3 and 25-hydroxyvitamin D3 in Ross 708 broilers subsequently challenged with coccidiosis: II. Immunological and inflammatory responses, and small intestine histomorphology

Authors: FATEMI, S - Mississippi State University; Collins Elliott, Katie; MACKLIN, K - Auburn University; BELLO, A - University Of Alberta; PEEBLES, E - Mississippi State University

Submitted to: Animals
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 4/12/2022
Publication Date: 4/14/2022
Citation: Fatemi, S.A., Collins Elliott, K.E., Macklin, K.S., Bello, A., Peebles, E.D. 2022. Effects of the in ovo injection of vitamin D3 and 25-hydroxyvitamin D3 in Ross 708 broilers subsequently challenged with coccidiosis: II. Immunological and inflammatory responses, and small intestine histomorphology. Animals. 12(8):1020. https://doi.org/10.3390/ani12081027.
DOI: https://doi.org/10.3390/ani12081027

Interpretive Summary: Coccidiosis is a parasite that often poses a problem for commercial meat-type chickens and causes economic impacts. The coccidia attack the bird’s intestine and diminishes the health of the bird as well as reduces the bird’s ability to effectively uptake nutrients from their feed. There are a few established methods to combat coccidiosis while raising chickens: including feeding vitamin D. This current study investigated how to better help defend the bird against coccidiosis by giving the unhatched chick within the egg vitamin D three days before the chick hatched. Giving a vaccine or other supplement is a commercial practice called in ovo vaccination or in ovo supplementation. This study thus investigated in ovo supplementation of vitamin D to help with a coccidiosis challenge by specifically looking at the inflammatory response of the birds and the appearance of the small intestine morphology. Birds were either not given an in ovo supplement of vitamin D (controls) or a dose of two different forms of vitamin D (D3 or 25OHD3) either separately or combined three days before they hatched. Vitamin D (D3) is naturally activated by two hydroxylation steps (adding a OH group) within the body: first at the liver to produce 25OHD3 and secondly at the kidney to produce the active form of vitamin D (1,25(OH)2D3). The hatched birds were raised, and the birds were challenged with coccidiosis at 2 weeks of age. Blood samples were collected from a subset of birds in each treatment at 2 and 4 weeks of age. Samples of the birds’ small intestines were also collected. The blood samples were analyzed for nitric oxide (NO) levels, which indicates the amount of an inflammatory response of the body and for IL-1ß levels, which indicates the immune response of the bird (pro-inflammatory cytokine) and may increase NO levels. Coccidiosis infections have been shown to increase the level of NO in the blood. The birds given 25OHD3 alone had a lowered NO level in their blood at 4 weeks of age. The birds given 25OHD3 alone in ovo also had a greater villus length to crypt depth ratio in their small intestine samples, which indicates a greater ability for nutrient/feed absorption within the small intestine. The IL-1ß levels did not differ between the treatments. Thus, giving meat-type chickens the 25OHD3 form of vitamin D in ovo may help reduce the inflammatory response and improve the intestine of birds combating a coccidiosis infection.

Technical Abstract: The effects of the in ovo administration of Vitamin D3 (D3) and 25-hydroxyvitamin D3 (25OHD3) on the small intestine morphology and inflammatory response of broilers challenged by coccidiosis were investigated. Live embryonated Ross 708 broiler hatching eggs were randomly assigned to one of the following 5 in ovo injection treatments at 18 d of incubation (doi): 1) non-injected; 2) diluent-injected; injected diluent containing either 3) 2.4 µg D3, 4) 2.4 µg 25OHD3, or 5) 2.4 µg D3 + 2.4 µg 25OHD3. A 50 µL solution volume was injected into each egg using an Inovoject multi-egg injector. At hatch, 4 male chicks were randomly assigned to each of 40 battery cages in each of 2 rooms, with 8 replicate cages per treatment group in each room. Only in ovo-injected treatments were challenged with a 20x live coccidial vaccine dosage at 14 d of age (doa). At 14 and 28 doa, blood samples were collected from 1 bird from each treatment-replicate cage in each room for determination of plasma IL-1ß and nitric oxide (NO) levels. From the same birds, spleen, bursa, liver, cecum, and small intestine weights, and 2 cm of the middle of the duodenum, jejunum, and ilium were excised for histomorphological analysis. Duodenum, jejunum, and ileum villus length (VL), crypt depth (CD), VL to CD ratio (RVC), and villus surface area (VSA) were measured. At 28 doa, plasma NO levels decreased (P=0.028) due to 25OHD3 alone when compared to all the other treatments. At 14 doa, in ovo injection of 25OHD3 alone resulted in higher duodenal RVC in comparison to all other treatments. At 28 doa, in ovo injection of 25OHD3 alone resulted in greater duodenal RVC in comparison to diluent, D3 alone, and the D3 + 25OHD3 combination treatments. These results indicate that the in ovo injection of 2.4 µg of 25OHD3 has the potential to improve the small intestine morphology and inflammatory response of broilers during a coccidiosis challenge.