
==== Front
Poult Sci
Poult Sci
Poultry Science
0032-5791
1525-3171
Elsevier

S0032-5791(24)00656-4
10.1016/j.psj.2024.104077
104077
IMMUNOLOGY, HEALTH AND DISEASE
Characterization of spray-dried plasma-induced immune response in Salmonella-challenged broiler chicks
Obanla Temitayo O. *
Khan Md Maruf *║
Adetunji Adedeji O. *¶
Ajuwon Kolapo M. †
Fahrenholz Adam C. ‡
Iwuozo Odinaka C. *
Adjei-Fremah Sarah §
Ekunseitan Deji A. *
Fasina Yewande O. yfasina@ncat.edu
*1
⁎ Department of Animal Sciences, North Carolina Agricultural and Technical State University, Greensboro, NC 27411, USA
† Department of Animal Sciences, Purdue University, West Lafayette, IN, 47907, USA
‡ Prestage Department of Poultry Science, North Carolina State University, Raleigh, NC 27695, USA
§ Department of Biological Sciences, Winston-Salem State University, Winston-Salem, NC 27110, USA
║ Department of Pathology and Laboratory Medicine and Legorreta Cancer Center, Brown University, Providence, RI 02903, USA
¶ Department of Agriculture, University of Arkansas at Pine Bluff, Pine Bluff, AR 71601, USA
1 Corresponding author: yfasina@ncat.edu
17 7 2024
11 2024
17 7 2024
103 11 10407712 4 2024
5 7 2024
© 2024 The Authors
2024
https://creativecommons.org/licenses/by-nc-nd/4.0/ This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
Spray-dried plasma (SDP) is a functional feed additive that has been established to improve performance and health of livestock. Understanding the effect of SDP in immune response modulation is essential to optimize its use for controlling Salmonella Enteritidis (SE) infection in chickens. This study was conducted to determine the levels of expression of selected cytokine genes in the ileum and cecal tonsil of SE-challenged broiler chicks. In a floor-pen housing, 320 broilers chicks were randomly assigned to 6 treatment groups: CX (unmedicated corn-soybean meal (SBM) basal without SDP), MX (unmedicated corn-SBM basal with antibiotic bacitracin methylene disalicylate (BMD) added at 0.055g/kg diet), PCX (unmedicated corn-SBM basal with SDP added at 30g/kg diet). Treatments SE, MSE, and PSE consisted of chicks inoculated with 7.46 × 108 CFU SE /mL at 1 d of age and given diets similar to CX, MX, and PCX, respectively. Samples of cecal tonsils and ileum were collected on d 3, 7 and 14 post infection for qRT-PCR analysis to determine the expression levels of interleukin (IL)-1β, interferon-γ (IFN-γ), IL-13, IL-17, IL-6, and transforming growth factor (TGF)-β genes. In the ceca tonsils, expression of IFN-γ was not affected by the interaction of Day and Treatment (P > 0.05). The level of the anti-inflammatory cytokine IL-13 was lower in MX and PCX on d 7 whereas high levels were expressed (P < 0.05) in MSE and PSE. In the ileum, expression of IL-17 and IFN-γ was significantly lower (P < 0.05) in PSE and MSE, but only PSE expressed lower IL-6 comparable to unchallenged treatments. On d 28 postchallenge, concentrations of anti-SE IgY and IL-6 protein were higher (P < 0.05) in the SE-challenged treatments compared to the unchallenged treatments. Overall, these results suggest that dietary SDP showed similar potency to BMD in modulating intestinal cytokine response against intestinal SE colonization in broiler chicks and therefore can be considered suitable alternative replacement for antibiotics in broiler production.

Key words

Spray-dried plasma
Salmonella Enteritidis
broiler chicken
immune response
intestinal cytokine
==== Body
pmcINTRODUCTION

Poultry is one of the essential components of agriculture saddled with the function of providing nutrition to the populace and contributing to the economy (Aljuwayd et al., 2023). The global shift from antibiotic growth promoters’ usage and consequent rise in antibiotic-free poultry production have heightened biosecurity issues, particularly due to the increase in intestinal disorders and pathologies, including both clinical and subclinical forms of destructive diseases such as necrotic enteritis and salmonellosis. Alternate options of control have been sought to replace the usage of conventional antibiotics and growth promoters and drive the move towards wholesome and residue-free poultry products.

The gastrointestinal system is a complicated environment that brings together a wide range of elements, including dietary biomaterials, pathogens, bacteria, peptides, and cellular concerns. An alteration in the system due to pathogenic or environmental challenge, heat stress, antinutritional factors etc., or the presence and interaction of these components determines the challenge put up against it by the system (Gomez-Osorio et al., 2021). The intestine (small and large) is a major component of the GIT and considered as the body's largest immunological organ (Dal Pont et al., 2023a). It has a complex inflammatory process that involves intestinal enterocytes present in the organ which secretes immune cells that migrate to the infection site. Intestinal inflammation can occur due to pathogen or environmental challenge, heat stress, antinutritional factors, and transportation stress. Although the use of antibiotics such as bacitracin methylene disalicylate (BMD) has been documented to influence birds’ performance, modify gut microbiota and create local anti-inflammatory effect (Aljuwayd et al., 2023) but the extensive negative impact occasioned by its attendant residue in the food chain system and the creation of resistant strains of foodborne pathogens such as salmonella necessitates the exploration of alternative strategies.

Salmonella Enteritidis (SE) is one of the leading causes of food borne illness around the world (Swaggerty et al., 2017; Acevedo-Villanueva et al., 2021). Gram negative bacteria are known to cause intestinal health problems and reduced growth performance in birds. It has been estimated that salmonellosis costs the United States Poultry industry approximately $11.6 billion annually (Wernicki et al., 2017). Antibiotics have been effective in the treatment of the disease; however, these compounds have not been able to penetrate through Salmonella-containing vacuole causing the bacteria to escape antimicrobial and phagocytic activities leading to public health and safety risks (Chen et al., 2019). It is established that the small intestine is the site for Salmonella invasion, however, the ceca has been implicated as the site for SE colonization and bacterial shedding (Cazals et al., 2022). SE has an outer membrane composed of rich lipopolysaccharide and its invasion of the chicken intestine induces an inflammatory process triggers the secretion of cytokines such as interleukin-1β (IL-1β), interleukin-6 (IL-6), and interferon-γ (IFN-γ) and other pro-inflammatory mediators such as chemokines by epithelial cells lining the intestine, ultimately leading to a severe inflammatory response (Kogut and Arsenault, 2017; Adetunji et al., 2020). In addition, it has been demonstrated that intestinal inflammation may induce the release of inflammatory cytokines (such as IL-6, IL-1β, TNF-α) that can cause disease in birds (Dal Pont et al., 2023b). The persistence of Salmonella in the intestinal tract of chickens is the main cause of disease propagation in poultry. A number of management and treatment approaches have been put forth to lessen the spread of Salmonella infection as an alternative to antibiotics in poultry production, one of which is the use of antimicrobial growth promoters including BMD ionophores and spray-dried plasma (SDP).

SDP was first proposed as a source of protein in piglets feed in late 1980 (Gatnau and Zimmerman, 1991), proving to have significant improvement in the performance of the piglets. It is a highly digestible protein derived from animal blood by separating plasma from whole blood by centrifugation (Blázquez et al., 2020; Blue et al., 2023). Unlike BMD, the SDP contains immunoglobulins, peptides, amino acids, enzymes, and lipid constituents which influence various immunological, biochemical, and metabolic processes to enhance overall wellbeing (Perez-Bosque et al., 2016). SDP has been reported to support intestinal health and broiler performances by enhancing digestive function and selectively modulate the microflora of microorganisms in the intestine (Zhang et al., 2015; Campbell et al., 2019) and reduce permeability modulating immune system (Wataha et al., 2013). SDP has been reported to improve growth performances, feed efficiency, and modulate immune response by reducing inflammations and improve intestinal health in broiler chickens (Campbell et al., 2019; Walter et al., 2019). In bacteria challenge studies, specific antibodies contained in SDP have important therapeutic and prophylactic value that act against enterotoxigenic Escherichia coli, leading to superior growth-enhancing effects (Owusu-Asiedu et al., 2002; Ruckman et al., 2020). Since SDP protects against Salmonella, its use as an alternative to antibiotics like BMD may be recommended (Boyer et al., 2015; Fasina et al., 2021) in maintaining performance and health of broiler chickens. Dietary SDP has been documented to act on mucosal systems to modulate systemic immunomodulation in addition to the gut immune response, through reduced expression of TNF-α, increased the expression values of TGF-β and IL-10, thereby restoring mucosal homeostasis and supporting anti-inflammatory effects (Pérez-Bosque et al., 2010; Pérez-Bosque et al., 2016; Campbell et al., 2019). Hence, replacement of antibiotics like BMD with an in-feed SDP will have significant potential in the poultry industry, offering a promising alternative to antibiotics by improving broiler chickens’ performance, intestinal health and ultimately consumer acceptance of poultry products.

Although several studies have focused on the functionality of SDP in improving feed efficiency, survivability, and weight gain (Campbell et al., 2019; Jababu et al., 2020, Fasina et al., 2021; Blue et al., 2023), its immunomodulatory role in the presence of Salmonella infection has not been explored. As a result, it was hypothesized that SDP may modulate and have direct anti-inflammatory effects on the intestine immune response. Therefore, this study was conducted to evaluate the effects of dietary addition of SDP on immune response in broiler chickens challenged with SE.

MATERIALS AND METHODS

The procedures used in this study were approved by the Institutional Animal Care and Use Committee (IACUC) of North Carolina Agricultural and Technical State University.

Experimental Animals and Design

A day-old Ross 708 male broiler chicks (n = 320) were obtained from a commercial hatchery and transported to the Poultry Research Unit of North Carolina Agricultural and Technical State University. On arrival, 20 chicks were randomly selected and euthanized by CO2 asphyxiation to confirm that the chicks were free of nalidixic acid resistant SE strain using a method previously described by Fasina et al. (2021). Chicks were weighed, and randomly assigned to 6 treatments in a completely randomized design. Treatment CX consisted of chicks fed unmedicated corn-soybean meal (SBM) basal without SDP. Treatment MX consisted of chicks given unmedicated corn-SBM basal into which BMD was added at 0.055 g/kg diet. Treatment PCX consisted of chicks given unmedicated corn-SBM basal into which SDP was added at 30 g/kg diet, as shown in the experimental diet (Table 1). Treatments SE, MSE, and PSE consisted of chicks that were given diets similar to CX, MX, and PCX, respectively, and were each inoculated with 1 mL of inoculum containing 7.46 × 108 CFU SE /mL at 1 d of age while the non-SE treatments were administered 1 mL of sterile BPW without Salmonella. Each of the 6 treatments consist of 5 replicate pens with 10 chicks per pen. The floor of each pen was covered with pine shavings. Experimental diets were formulated to meet the recommendations of the National Research Council (1994), and chicks were allowed access to feed and water ad libitum throughout the experiment that lasted for 28 d as previously described by Fasina et al. (2021). Growth performance parameters such as feed intake, body weight and mortality were monitored throughout the experiment and FCR was calculated at d 28.Table 1 Experimental diets composition1 (% “as is”).

Table 1	Starter Diets1 (d 1–14)	Grower Diets1 (d 15–28)	
Ingredient	Control diet	BMD diet	SDP diet	Control diet	BMD diet	SDP diet	
Corn (7.5% Crude protein)	51.46	51.45	55.94	56.77	56.76	61.24	
Soybean meal (47.5% Crude Protein)	40.39	40.40	35.06	35.43	35.43	30.09	
Spray-dried plasma (SDP, AP920)	—	—	3.00	—	—	3.00	
Poultry fat	3.64	3.65	1.96	4.00	4.00	2.31	
Limestone	1.07	1.07	1.19	0.64	0.64	0.76	
Mono-Dicalcium phosphate	2.03	2.03	1.85	1.84	1.84	1.67	
Salt NaCl	0.40	0.40	0.23	0.40	0.40	0.24	
Sodium bicarbonate	0.02	0.02	—	0.02	0.02	—	
L-Lysine HCl 98%	0.13	0.13	0.05	0.11	0.11	0.03	
DL-Methionine 99.0%	0.34	0.34	0.28	0.30	0.30	0.24	
L-Threonine 98.5%	0.11	0.11	0.04	0.09	0.09	0.02	
NCSU poultry vitamin premix2	0.05	0.05	0.05	0.05	0.05	0.05	
NCSU poultry mineral premix3	0.20	0.20	0.20	0.20	0.20	0.20	
Bacitracin (Antibiotic, g/kg)	—	0.055	—	—	0.055	—	
Choline chloride 60%	0.10	0.10	0.10	0.10	0.10	0.10	
Selenium Premix	0.05	0.05	0.05	0.05	0.05	0.05	
Analyzed nutrient composition4							
 Metabolizable energy (Kcal/kg)	3,117	3,131	3,137	3,283	3,212	3,234	
 Crude Protein, %	24.63	24.56	24.06	18.81	23.94	22.25	
 Crude Fat, %	4.74	5.03	4.18	5.06	5.84	5.59	
 Crude Fiber, %	2.3	2.4	2.3	2.2	2.4	2.2	
 Ash, %	6.32	6.15	5.35	4.59	5.68	5.19	
Calculated nutrient composition							
 Total Sulfur Amino Acids, %	1.03	1.03	1.03	0.95	0.95	0.94	
 Lysine, %	1.42	1.42	1.41	1.27	1.27	1.27	
 Calcium, %	0.96	0.96	0.96	0.75	0.75	0.75	
 Available phosphorus, %	0.48	0.48	0.48	0.44	0.44	0.44	
1 Diets used in this study included the following: i) unmedicated corn-soybean meal (SBM) basal without SDP (Control diet); ii) unmedicated corn-SBM basal into which bacitracin methylene disalicylate (BMD) was added at 0.055g/kg diet (BMD diet); and iii) SDP diet in which spray-dried plasma was incorporated into unmedicated corn-SBM basal at 3% level (i.e. 30 g/kg diet). Each of these 2 diets were separately formulated for the starter (d 1 to 14), and Grower (d 15 to 28), phases of broiler production cycle.

2 Vitamin Premix, supplied per kilogram of diet: Vitamin A (6,600 IU), Vitamin D (1,980 IU), Vitamin E (33 IU), Vitamin B12 (0.02 mg), Biotin (0.13 mg), Menadione (1.98 mg), Thiamine (1.98 mg), Riboflavin (6.60 mg), d-Pantothenic Acid (11.0 mg), Vitamin B6 (3.96 mg), Niacin (55.0 mg), Folic Acid (1.1 mg).

3 Mineral Premix, supplied per kilogram of diet: Manganese (Mn), 60 mg; Zinc (Zn), 60 mg; Iron (Fe), 40 mg; Copper (Cu), 5 mg; Iodine (I), 1.2 mg; Cobalt (Co), 0.5 mg.

4 Experimental diets were analyzed for proximate nutrient composition by Eurofins Scientific Inc. Nutrient Analysis Center, 2200 Rittenhouse Street, Suite 150, Des Moines, IA 50321.

Preparation of Salmonella Inoculum, and Challenge

Frozen stock culture of SE (strain type) was briefly thawed and a volume of 10 µl was collected and inoculated into a tube containing 10 ml of Tryptic Soy Broth (TSB, MP Biomedicals, Irvine, CA). The broth containing SE bacteria was incubated overnight at 37°C (Thermo Scientific Heratherm Advanced Protocol Microbiological Incubator, Waltham, MA) and streaked on Xylose Lysine Tergitol 4 (XLT4, MP Biomedicals, Irvine, CA) agar plate to which 0.1% nalidixic acid solution (50 µg/mL) had been added. The plates that were streaked were then incubated for 48 h at 37°C. Thereafter, black presumptive colonies of SE were collected and inoculated into a tube containing 10 ml of sterile TSB and incubated at 37°C for 24 h. The resulting culture was used to prepare the inoculum for chick challenge for the SE-challenge treatments while 1 mL of sterile BPW was administered to the non-SE challenge treatments as previously described by Fasina et al. (2021).

Sample collection

On the d 3, 7, and 14, 2 birds/pen were randomly selected from each treatment and ileum and cecal tonsils were aseptically removed and stored at −80°C for subsequent analysis of the expression levels of inflammatory-related cytokine genes. Furthermore, on d 28, blood was collected from the brachial (wing) vein with the aid of a sterile 23 gauge 1′' needle attached to sterile EDTA vacutainer tubes. Thereafter, the blood collected was centrifuged at 1,500 x g for a duration of 10 min, and the supernatant was carefully collected to obtain plasma and stored at −80°C.

RNA Extraction, cDNA Synthesis and RT-PCR Analyses

Total RNA from ileum tissue and cecal tonsils (30 mg/sample) were extracted after homogenization with a MiniBeadbeater-96TM (Biospec Products, Inc., Bartlesville, OK) using RNeasy mini kit (Qiagen, Waltham, MA) following the manufacturer's protocol. The RNA quality was determined spectrophotometrically by A260/A280 ratio. Thereafter, 1 µg of total RNA was reverse transcribed into complementary DNA (cDNA) using the iScript Reverse Transcription Supermix kit (Bio-Rad, Hercules, CA) based on manufacturer's instructions. The expression levels of genes involved in immune response were measured in ileum tissue and cecal tonsils by real-time polymerase chain reaction using CFX connect Real-Time PCR Detection System (Bio-Rad, Hercules, CA) using SsoAdvanced Universal SYBR Green Supermix (Bio-Rad, Hercules, CA). The volume of total reaction mix was 20 µL, including 10 µL SsoAdvanced Universal SYBR Green Supermix (2x), 1 µL of each primers pair and 3 µL cDNA template. The PCR procedures were 95°C for 3 min, followed by 40 cycles of 95°C for 5 s, annealing temperature for 30 s and 72°C for 10 min. To normalize RNA levels between samples within an experiment, the mean threshold cycle (Ct) values for the amplification products were calculated by pooling values from all samples in that experiment. Each sample was in triplicate, and the expression levels of genes were calculated by 2−△△Ct method (Livak and Schmittgen, 2001). According to Chauhan et al. (2021) classification, the cytokines evaluated are proinflammatory, interleukin 1 beta (IL-1β); Th1, interferon gamma (IFN-ϒ); Th2, interleukin 6 and 13 (IL-6, IL-13); Th17, interleukin 17 (IL-17) and anti-inflammatory transforming growth factor beta (TGF-β). Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) was considered a housekeeping gene. Primers used for RT-PCR of the cytokines were listed in Table 2.Table 2 List of selected primers for quantitative RT-PCR assay

Table 2Class	Target gene	Primer sequence (5′ – 3′)	Reference	
House keeping	Glyceraldehyde 3-phosphate dehydrogenase (GAPDH)	F: 5′-GGTGGTGCTAAGCGTGTTAT-3′
R: 5′-ACCTCTGTCATCTCTCCACA-3′	Fasina and Lillehoj, 2019	
Proinflammatory	Interleukin 1 beta (IL-1β)	F: 5′-TGGGCATCAAGGGCTACA-3′
R: 5′-TCGGGTTGGTTGGTGATG-3′	Fasina and Lillehoj, 2019	
Th1	Interferon Gama (IFN-y)	F:5′-AGCTGACGGTGGACCTATTATT-3′
R: 5′-GGCTTTGCGCTGGATTC-3′	Fasina and Lillehoj, 2019	
Th2	Interleukin 6 (IL-6)	F: 5′-CTCGTCCGGAACAACCTCAA-3′
R: 5′-GGAGAGCTTCGTCAGGCATT-3′	Designed using NCBI Primer-BLAST Software	
Interleukin 13 (IL-13)	F: 5′ -CCAGGGCATCCAGAAGC-3′
R: 5′-CAGTGCCGGCAAGAAGTT -3′	Fasina and Lillehoj, 2019	
Th17	Interleukin 17 (IL-17)	F: 5′-CTCCGATCCCTTATTCTCCTC-3′
R: 5′-AAGCGGTTGTGGTCCTCAT-3′	Fasina and Lillehoj, 2019	
Anti-inflammatory	Transforming growth factor beta (TGF- β)	F: 5′-CGGGACGGATGAGAAGAAC-3′
R: 5′-CGGCCCACGTAGTAAATGAT-3′	Fasina and Lillehoj, 2019	

Enzyme-Linked Immunosorbent Assay

On d 28, the concentration of IL-6 and anti-SE IgY proteins in the plasma of chicks were detected according to instructions of the Enzyme-linked immunosorbent assay (ELISA) kit for IL-6 (ECH6RB, Thermo scientific, Waltham, MA) and anti-SE IgY (DEIABL37, Creative Diagnostic, Shirley, NY). Briefly, a microtiter plate reader; Synergy H1 (Biotek, Winooski, VT) was used to measure the absorbance at 450 nm and a 4-parameter logistic curve fit (standard curve) and expressed in nanograms per millimeter (ng/mL).

Statistical Analysis

All quantitative RT-PCR cytokine gene expression data, antibody ELISA data, and growth performance were subjected to 1-way ANOVA using the general linear models (GLM) procedure of SAS (SAS Inc., 2004) except for ceca tonsil d 3, 7 and 14 data that was subjected to 3 × 6 (IL-17, IL-1β, IL-6) and 2 × 6 (IFN-Ƴ, IL-13, TGF-β) factorial analysis. Multiple comparisons were analyzed using ANOVA followed by Tukey test. The P < 0.05 was considered to be statistically significant. Data was displayed as mean ± SEM.

RESULTS

Growth Performance Parameters and SE Infection Confirmation

Overall growth performance from d 1 to d 28 (Table 3) shows that PCX and PSE had similar body weight, and CX, SE and MSE also had similar body weight (P > 0.05), however, MX had highest (P < 0.05) body weight (BW) of 1.603 kg/ bird. The BMD supplementation increased (P < 0.05) the body weight gain (BWG) in SE-challenged and unchallenged treatments, however, statistically similar values (P > 0.05) of BWG were recorded in all other challenged and unchallenged treatment groups. In the treatment without SE-challenge, CX had higher feed conversion ratio (FCR) values (0.715; P < 0.05), MX and PCX had similar values compared to SE-challenged treatments, whereas PSE had least FCR values (0.691) and MSE had highest FCR value (0.973) in the SE-challenged treatments. Additionally, SE had highest (P < 0.05) feed intake (FI) of 2.035 kg/bird followed by CX and MX, 1.843 and 1.828 respectively, but other treatments recorded similar (P > 0.05) values for FI. The mortality percentage (P > 0.05) was similar across all treatment groups (P = 0.391) with or without SE challenge.Table 3 Effect of dietary spray-dried plasma on growth performance of broiler chickens (d 1–28).

Table 3	(Parameters measured)2	
Treatment1	Body weight (BW, kg/bird)3	Body weight gain (BWG, kg/bird)	FCR4 (kg:kg)	Feed intake (FI, kg/bird)	Mortality (%)	
CX	1.458bc	1.000c	0.715b	1.843b	3.34	
MX	1.603a	1.133b	0.680de	1.828bc	1.67	
PCX	1.355c	1.035bc	0.670e	1.720d	0.00	
SE	1.576ab	1.063bc	0.706bc	2.035a	6.67	
MSE	1.508ab	1.468a	0.973a	1.760cd	5.00	
PSE	1.355c	0.970c	0.691cd	1.785bcd	5.00	
P- value	0.0015	<0.0001	<0.0001	<0.0001	0.42	
a,b,c,d,e Mean values bearing different superscript letters within a column are significantly different (P < 0.05).

1 Treatment CX consisted of chicks fed unmedicated corn-soybean meal (SBM) basal without SDP; Treatment MX consisted of chicks given unmedicated corn-SBM basal into which bacitracin methylene disalicylate (BMD) was added at 0.055 g/kg diet; Treatment PCX consisted of chicks given unmedicated corn-SBM basal into which SDP was added at 30g/kg diet; Treatments SE, MSE, and PSE, consisted of chicks that were given diets similar to CX, MX, and PCX, respectively, and were each inoculated with 7.48 × 108 CFU Salmonella Enteritidis /mL at 1 d of age.

2 Values represent the mean of 4 replicate pens per treatment.

3 Values are based only on weight of live birds.

4 FCR = feed conversion ratio calculated as feed-to-gain ratio and adjusted for mortality by including the gains of dead birds in the calculations.

Establishment of SE infection was confirmed, with the treatment groups challenged with SE on d 3 showing that BMD and SDP-fed chicks had similar (P > 0.05) cecal SE (3.39 log 10 CFU / g and 3.58 log 10 CFU / g, respectively), but these levels were lower (P < 0.05) than that of CON-fed chicks (5.68 log 10 CFU /g). A similar trend was observed on d 7 and 14. Results showed that SE infection was successfully established in chicks assigned to SE, MSE, and PSE treatments (Table 4).Table 4 Effect of spray-dried plasma on the concentration of Salmonella Enteritidis in ceca of broiler chickens.

Table 4	Log10 CFU/g cecal contents	
Treatment	d 3	d 7	d 14	
SE	5.68 ± 0.36a	4.17 ± 0.18a	2.48 ± 0.05a	
MSE	3.39 ± 0.14b	2.41 ± 0.12b	1.53 ± 0.09b	
PSE	3.58 ± 0.11b	2.26 ± 0.09b	1.06 ± 0.05b	
P- value	<0.0001	<0.0001	<0.0001	
a-b Mean values bearing different superscript letters within a column are significantly different (P < 0.05).

Gene Expression of Intestinal Cytokines

In the ceca tonsils, IL-17 expression level was not affected by the Day, Treatment, and the interaction of Day x Treatment (P > 0.05) (Figure 1A). Figure 1B also shows that IFN-γ expression level was not affected by Day x Treatment (P > 0.05). However, PSE had the highest level of expression (P < 0.05) compared to other treatment groups. Furthermore, IL-1β expression was significantly affected (P < 0.05) by the interaction of Day x Treatment (Figure 1C). However, IL-1β expression among the challenged treatments was lower on d 14, while the unchallenged treatments were not significantly different across all the days. Furthermore, IL-6 gene expression was influenced by the interaction of Day x Treatment (P < 0.05). There was an increase in the expression of IL-6 among the challenged treatments on d 7 compared to unchallenged birds, but on d 14 this had declined (Figure 1D). In addition, there was a significant (P < 0.05) Day x Treatment interaction in the expression of IL-13 such that there was upregulation of the cytokine in PSE compared to other treatments on d 3 and d 7 (Figure 1E). There was also a Day x Treatment interaction effect (P < 0.05) in the expression of TGF-β gene (Figure 1F) such that PSE is upregulated on d 7 while other treatments were unchanged on both days.Figure 1 Expression of (A) IL-17 (B) IFN-γ (C) IL-1β (D) IL-6 (E) IL-13 (F) TGF-β genes in the cecal tonsils of unchallenged and SE-challenged broiler chickens on different days. The data are expressed as means ± SEM, with n = 10 per treatment; a-g means without a common superscript are different. Statistically significant differences are indicated with P < 0.05.

Figure 1

In the ileum, least IL-6 gene was expressed in PSE treatment (P < 0.05) compared to the other SE-challenged treatments, which is not statistically different (P > 0.05) from the unchallenged treatments except CX (P < 0.05); IFN-Ƴ had similar expression in all treatments except PCX, while statistically similar values were recorded in IL-17 for PSE and MSE and other unchallenged treatments (Figure 2A, B and D). However, there was no significant difference (P > 0.05) in the expression of IL-1β among treatments (Figure 2C).Figure 2 Expression of (A) IL-17 (B) IFN-γ (C) IL-1β (D) IL-6 genes in the ileum of unchallenged and SE-challenged broiler chickens on d 7. The data are expressed as means ± SEM, with n = 10 per treatment; a, b, c means without a common superscript are different. Statistically significant differences are indicated with P < 0.05.

Figure 2

Effects of Dietary Supplementation of SDP on Plasma Concentrations

Furthermore, for IL-6 protein and anti-SE IgY concentrations, the SE-challenged treatments had a higher concentration (P < 0.05) for both proteins compared to the unchallenged treatments (Figures 3A and 3B).Figure 3 Concentration of (A) Anti-Salmonella IgY and, (A) IL6 IgY in the plasma of unchallenged and SE-challenged broiler chickens on d 28. The data are expressed as means ± SEM, with n = 10 per treatment; a, b, c means without a common superscript are different. Statistically significant differences are indicated with P < 0.05.

Figure 3

DISCUSSION

In this study, the effect of dietary SDP supplementation in broiler chickens challenged with SE was evaluated. Dietary supplementation of SDP has been shown to have beneficial effects on the growth performance and intestinal health in broiler chickens (Beski et al., 2015; Campbell et al., 2019; Jababu et al., 2020; Blue et al., 2023). In the present study it was observed that SDP positively influenced the FCR in both challenged and unchallenged groups. Accordingly, similar efficacy of dietary SDP was reported by Henn et al (2013) with lowered feed intake and improved FCR, thus, this may be attributed to the fact that SDP improved the jejunal goblet density and maintained the intestinal villi regeneration (Jababu et al., 2020), and hence the development of the small intestinal mucosa leading to better absorption and utilization of nutrients for biological activities, including growth (Campbell et al., 2019). Dietary SDP reduced the BW and BWG, while least feed intake was observed to initiate optimum performance in this study, this is in contrast to studies by Walters et al. (2019) and Daneshmand et al. (2023) who reported increased body weight when dietary SDP was fed beyond 28 d. The reduced BW and BWG may be attributed to the dosage (30 g/kg) and duration (28 d) of this study, unlike the reported dosage by Campbell et al. (2019) and Jababu et al. (2020) respectively. Moreover, increased nutrients utilization is known to be responsible in the activation of several immune systems which play vital roles in the stabilization of immune homeostasis and structural functions (Campbell et al., 2019) of the intestines to harness absorbed nutrients into better growth performance. Therefore, the reduced BW and BWG may be related to the diversion of the absorbed nutrients in the activation of immune functions compared to production functions. The mortality percentage reported in this study was similar across Dietary treatment x SE challenge, it was however numerically higher in the SE challenged groups compared to the nonchallenged group. This is uncommon as SE challenged chicks have been reported to have mortality percentage ranging from 0 - 15% (Shittu et al., 2014; Fasina et al., 2021) or more (Amer et. al., 2020) depending on the virulence of Salmonella, age of bird and physiological stressors (Shaji, et al., 2023; Orimaye et al., 2024).

A successful infection of SE was established in the ceca of the challenged chicks; thus, SDP and BMD were effective in reducing ceca Salmonella colonization during the first 14 d. The SDP reduction of intestinal SE may be attributed to the immunoglobulins and other immune modulating ability of SDP, which improved the structure of intestinal mucosa barrier by inhibiting the spread of the pathogens and reducing the localized inflammation (Perez-Bosque et al., 2016) caused by Salmonella infection, thus enhance the production of goblet cells in the gut epithelium (Liu et al., 2018) and subsequent mucin production (Jababu et al., 2020), which is reported to inhibit pathogenic bacteria adhesion to the gut (Johansson and Hansson, 2016) and reduce subsequent risk of subclinical infection to other vital organs (Balan et al., 2021). Inclusion of in-feed SDP suppressed SE proliferation just like BMD by inhibiting the cell wall biosynthesis leading pathogen death and growth retardation (Hutchings et al., 2019).

The mucosal immune system which consists of the gut-associated lymphoid tissue (GALT) can be found in the entire intestinal lumen and it is essential in protecting the epithelial cells from stressors and pathogenic microorganisms (Kiyono and Fukuyama, 2004; Granger et al., 2006). In most cases, the mucosal immune response is triggered by the invasion of pathogen which culminates in the release of pro-inflammatory cytokines (Petersen et al., 2004; Perez-Bosque et al., 2010). Overall, the innate immune system plays a vital role in host defense against infection. Moreover, in this study it was demonstrated that SDP and BMD dietary supplementations influence the levels of cytokine gene expression and their protein concentration in the gut of broiler chicken. Cytokines such as IL-1β and IL-6 exhibit pro-inflammatory properties and are involved in innate immunity. Specifically, interleukin 6 (IL-6) is implicated in the recruitment of immune cells to the predilection site as well as in the transition from innate to acquired humoral immunity (Kaiser et al., 2000). Our results showed that in ceca tonsils, the group without SE-challenge, PCX and MX treatments did not alter IL-17 and IL-6 expression compared to the control. Immunoglobulins account for about 20 percent of SDP dry weight and play a key role in SDP action against intestinal pathogens (Pierce et al., 2005; Ferreira et al., 2009;). Previously, a study showed that oral administration of SDP enhanced immune response as a result of its high immunoglobulin content (Ulfman et al., 2018). Similarly, the SE challenged increased serum levels, due to pro-inflammatory activities of IL-6 and IL-1β to trigger antibody productions, suggesting that SE may induce inflammatory response (Liu et al., 2019). In this present study, the ceca tonsils among the SE-challenged treatments, PSE treatment increased the level of expression for IL-6 and IL-1β (pro-inflammatory cytokines) as well as TGF-β and IL-13 (anti-inflammatory cytokines) compared to SE on d 7 which were not altered by PCX and MX treatments. Interleukin-1β (IL-1β) has been described as a potent pro-inflammatory cytokine that is crucial for host defense responses to infection. The preventive effects of plasma supplementation action on gut inflammation are characterized by the modulation of intestinal cytokines which causes an increase in the expression level of anti-inflammatory cytokines (Pérez-Bosque et al., 2010). In contrast, we did not observe any elevation in the expression of anti-inflammatory cytokines among the unchallenged treatments in this present study, suggesting the absence of intestinal inflammation. However, the surge in the expression of pro-inflammatory cytokines in the PSE treatment ultimately regulated the inflammation resulting from SE challenge, which in turn triggered a compensatory anti-inflammatory reaction (Jaffer et al., 2010) to cushion the pathogenic effects. Although the interaction of treatment and day had no effect on the IFN-γ, the treatment however had distinct elicited differed responses with the BMD reducing the expression of adaptive cytokines compared to upregulation by SDP. BMD has been opined by to reduce the mRNA expression of the adaptive cytokines IFN-γ, IL-10 and IL-4 and in the ceca of broilers at early stages of growth (7 d and 14 d) and pro-inflammatory T-cell cytokines (IL-17, IL-2 and IFN-γ) in the jejunum (Kumar et al., 2018; Dal Pont et al., 2023b).

In broiler chickens, the small intestine is the major site of nutrient utilization and metabolism, it also plays an active role in immunological functions that ultimately influence the overall health status of the animals. Primarily, ileum is composed of a large proportion of GALT, which affects animal health by providing nutrients, protection against pathogens and immune system development (Hong et al., 2021). The results in the ileum revealed that without SE-challenge, PCX and MX treatments also did not alter IL-17 and IL-6 expression compared to the control, however, PCX treatment increased the expression of IFN-γ in the ileum. We speculate that the increase in IFN-γ in the PCX treatment in the ileum was because SDP triggered the early expression of IFN-γ as a result of its higher immunoglobulin content in a bid to prime effector immune cells (Rodríguez et al., 2023). However, SE treatment increased the expression of IL-17 and IL-6 while PSE and MSE decreased the release of these pro-inflammatory cytokine genes. Previous challenge studies using pigs have also shown that SDP supplementation decreases the level of IL-6 compared to treatments without SDP (Touchette et al., 2002). Similar values were observed for the expression of IFN-γ for all SE-challenged treatments, this may be attributed to IFN-γ’s early activation of effector immune cells contributed by the dietary effect of BMD and SDP, which is crucial for early control of pathogen spread (Alspach et al., 2019), as they act by controlling the response of the cytokine of adaptive and innate immune system, hence reduce intestinal induced inflammation (Dal Pont et al., 2023b). In the early stages of infection, phagocytes are activated by the inflammation of the affected tissues. According to Iwasaki and Medzhitov (2010), SE colonization correlate with the expression of the cell mediated cytokine to recruit macrophages and monocytes involved in the activation of innate immunity. Also, large amounts of IFN-γ are produced from mainly NK cells, to promote adaptive immunity. Furthermore, the result of this study of IFN-γ expression agrees with Perez-Bosque et al. (2008), who stated that SDP modulated the immune response of rats administered Staphylococcus aureus toxin by inhibiting the effect of the toxin on the release of pro-inflammatory cytokines (IL-1β, IL-6, IFN-γ, etc.) while enhancing the expression of anti-inflammatory cytokines (IL-4, IL-10, IL-11, IL-13, IL-6, TGF-β, etc.) in the gut. The expression of IL-1β in the ileum tissue was similar for both challenged and unchallenged group, contrary to SDP supplementation reported to reduce the expression of TNF-α, and IL-1β in ileum tissues (Miska et al., 2022). The disparity between our result in the ileum on d 7, may be attributed to differences in the tissue location and function as well as the distribution of NK and T cells (Göbel et al., 2001).

Previously studies showed that dietary SDP supplementation decreased IL-6 level compared to other challenged groups (Daneshmand et al., 2023; Touchette et al., 2002; Pérez-Bosque et al., 2008). Contrarily, the findings of this present study disagree with the previous studies, as the concentration of IL-6 in the PSE treatment increased similar to the SE treatment. Additionally, this study observed that the concentration of anti-SE IgY level in the PSE treatment also increased similar to IL-6. The supplementation of dietary SDP did not influence the expression of pro-inflammatory cytokine (IL-6) in rats (Pérez-Bosque et al., 2010), this suggests that SDP acts locally and does not play a vital role in preventing the spread of SE infection in the broiler chickens through the systemic blood circulation. Overall, by reducing intestinal inflammation, SDP may be preventing the spread of bacteria or other antigens, hereby reducing immune activation locally (Dal Pont et al., 2023b).

In conclusion, performance data from the present study demonstrates that BWG and BW were improved in BMD compared to SDP in both SE challenged and unchallenged scenarios while SDP achieved the best feed efficiency. Our study demonstrates that SDP modulates immune response by decreasing intestinal expression of inflammatory genes in SE challenged broiler chicken. It provides evidence for the decrease of cytokine pro-inflammatory genes in localized intestinal tissues, SDP did not cause a similar reduction in the plasma to promote the spread of SE infection at the systemic level. These results suggest that dietary SDP showed similar potency to BMD in modulating intestinal cytokine response against intestinal SE colonization in broiler chickens.

DISCLOSURES

The authors of this manuscript hereby declare no conflict of interest. Manuscript has been “spell checked” and “grammar checked.” All references mentioned in the Reference List are cited in the text, and vice versa. Permission has been obtained for use of copyrighted material.

ACKNOWLEDGMENTS

This study was supported by the National Institute for Food and Agriculture of the United States Department of Agriculture, Evans Allen Project No. NC.X-329-5-20-120-1, in the Agricultural Research Program, North Carolina Agricultural and Technical State University. The authors also thank members of the Poultry Ethnomedicine Research Laboratory at North Carolina Agricultural and Technical State University (Greensboro, NC) for their technical support.
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REFERENCES

Acevedo-Villanueva K.Y. Renu S. Shanmugasundaram R. Akerele G.O. Gourapura R.J. Selvaraj R.K. Salmonella chitosan nanoparticle vaccine administration is protective against Salmonella Enteritidis in broiler birds PLoS ONE 16 2021 e0259334
Adetunji A.O. Kawai T. Shimada M. Impact of lipopolysaccharide administration on luteinizing hormone/choriogonadotropin receptor Lhcgr expression in mouse ovaries J. Reprod. Immu. 142 2020 103193
Aljuwayd M. Malli I.A. Kwon Y.M. Application of eugenol in poultry to control salmonella colonization and spread Vet. Sci. 10 2023 151 36851455
Alspach E. Lussier D.M. Schreiber R.D. Interferon γ and its important roles in promoting and inhibiting spontaneous and therapeutic cancer immunity Cold Spring Harbor Per. Bio 113 2019 a028480
Amer M.M. Amer A.M. Hassan E.R. Ghetas A.M. Salmonella enteritidis in broiler chickens: Isolation, antibiotic resistance phenotyping and efficacy of colistin on control of experimental infection Int. J. Vet. Sci 9 2020 267 272
Balan P. Staincliffe M. Moughan P.J. Effects of spray-dried animal plasma on the growth performance of weaned piglets—a review J. Anim. Physio. Anim. Nutr. 105 2021 699 714
Beski S.S.M. Swick R.A. Iji P.A. Specialized protein products in broiler chicken nutrition: A review Anim. Nutr. 1 2015 47 53 29766993
Blázquez E. Rodríguez C. Ródenas J. Segalés J. Pujols J. Polo J. Biosafety steps in the manufacturing process of spray-dried plasma: A review with emphasis on the use of ultraviolet irradiation as a redundant biosafety procedure Por. Hth. Mgt. 6 2020 16
Blue C.E.C. Jababu Y. Ibrahim S.A. Minor R.C. Williams L.L. Adetunji A.O. Ali R. Young L.S. Fasina Y.O. Spray-dried plasma promotes broiler chick growth by enhancing immune surveillance Anims 13 2023 1436
Boyer P.E. D'Costa S. Edwards L.L. Milloway M. Susick E. Borst L.B. Thakur S. Campbell J.M. Crenshaw J.D. Polo J. Moeser A.J. Early-life dietary spray-dried plasma influences immunological and intestinal injury responses to later-life Salmonellatyphimurium challenge Br. J. Nutr. 113 2015 783 793 25671331
Campbell J.M. Crenshaw J.D. González-Esquerra R. Polo J. Impact of spray-dried plasma on intestinal health and broiler performance Micro 7 2019 219
Cazals A. Estellé J. Bruneau N. Coville J.L. Menanteau P. Rossignol M.N. Jardet D. Bevilacqua C. Rau A. Bed'Hom B. Velge P. Calenge F. Differences in caecal microbiota composition and Salmonella carriage between experimentally infected inbred lines of chickens Gen. Sel. Evo. 541 2022 1 15
Chen J. Ying G.G. Deng W.J. Antibiotic residues in food: extraction, analysis, and human health concerns J. Agric. Food Chem. 67 2019 7569 7586 31198037
Dal Pont G.C. Lee A. Bortoluzzi C. Farnell Y.Z. Gougoulias C. Kogut M.H. Novel model for chronic intestinal inflammation in chickens: (2) immunologic mechanism behind the inflammatory response Devpt. Comp. Immu. 138 2023 104524
Dal Pont G.C. Lee A. Bortoluzzi C. Farnell Y.Z. Gougoulias C. Kogut M.H. Bacitracin supplementation as a growth promoter down-regulates innate and adaptive cytokines in broilers’ intestines Poult 2 2023 411 417
Daneshmand A. Sharma N.K. Dao T.H. Barekatain R. Swick R.A. Wu S.B. Spray-dried porcine plasma enhances feed efficiency, intestinal integrity, and immune response of broilers challenged with necrotic enteritis Poult. Sci. 102 2023 102431
Fasina Y.O. Lillehoj H.S. Characterization of intestinal immune response to Clostridium perfringens infection in broiler chickens Poult. Sci. 98 2019 188 198 30239962
Fasina Y.O. Obanla T.O. Ferket P.R. Shah D.H. Comparative efficacy of spray-dried plasma and bacitracin methylene disalicylate in reducing cecal colonization by Salmonella Enteritidis in broiler chickens Poult. Sci. 100 2021 101134
Ferreira A.S. Barbosa F.F. Tokach M.D. Santos M. Spray-dried plasma for pigs weaned at different ages Rec. Pat. Food Nutri. Agric. 13 2009 231 235
Gatnau R. Zimmerman D.R. Spray dried porcine plasma (SDPP) as a source of protein for weanling pigs in two environments J. Anim. Sci. 69 1991 103
Göbel T.W. Kaspers B. Stangassinger M. NK and T cells constitute two major, functionally distinct intestinal epithelial lymphocyte subsets in the chicken Intl. Immu. 136 2001 757 762
Gomez-Osorio L.M. Yepes-Medina V. Ballou A. Parini M. Angel R. Short and medium chain fatty acids and their derivatives as a natural strategy in the control of necrotic enteritis and microbial homeostasis in broiler chickens Front. Vet. Sci. 8 2021 773372
Granger D.N. Grisham M.B. Kevil C.G. Recruitment of inflammatory and immune cells in the gut: physiology and pathophysiology In: Physiology of the gastrointestinal tract 2006 Academic Press Cambridge, MA 1137 1162
Henn J.D. Bockor L. Vieira M.S. Ribeiro A.M.L. Kessler A.M. Albino L. Crenshaw J.D. Rostagno H. Campbell J.M. Rangel L.F.S. Inclusion of porcine spray-dried plasma in broiler diets J. Appl. Poult. Res. 22 2013 229 237
Hong Y. Cheng Y. Guan L. Zhou Z. Li X. Shi D. Xiao Y. Bacillus amyloliquefaciens TL downregulates the ileal expression of genes involved in immune responses in broiler chickens to improve growth performance Micro. 19 2021 382
Hutchings M.I. Truman A.W. Wilkinson B. Antibiotics: Past, present and future Cur. Opinio. Micro 51 2019 72 80
Iwasaki A. Medzhitov R. Regulation of adaptive immunity by the innate immune system Sci 327 2010 291 295
Jababu Y. Blue C. Ferket P.R. Fasina Y.O. Comparative effects of Spray-dried plasma and bacitracin methylene disalicylate on intestinal development in broiler chicks Intl. J. Poult. Sci. 19 2020 161 168
Jaffer U. Wade R.G. Gourlay T. Cytokines in the systemic inflammatory response syndrome: A review HSR Proc. Inten. Care Cardiovasc. Anesth. 2 2010 161 175
Johansson M.E.V. Hansson G.C. Immunological aspects of intestinal mucus and mucins Nat. Rev. Immunol. 16 2016 639 649 27498766
Kaiser P. Rothwell L. Galyov E.E. Barrow P.A. Burnside J. Wigley P. Differential cytokine expression in avian cells in response to invasion by Salmonella typhimurium, Salmonella Enteritidis and Salmonella gallinarum Microb 146 2000 3217 3226
Kiyono H. Fukuyama S. NALT-versus Peyer's-patch-mediated mucosal immunity Nat. Rev. Immuno. 4 2004 699 710
Kogut M.H. Arsenault R.J. Immunometabolic phenotype alterations associated with the induction of disease tolerance and persistent asymptomatic infection of Salmonella in the chicken intestine Front. Immunol. 8 2017 252032
Kumar S. Chen C. Indugu N. Werlang G.O. Singh M. Kim W.K. Thippareddi H. Effect of antibiotic withdrawal in feed on chicken gut microbial dynamics, immunity, growth performance and prevalence of foodborne pathogens PLoS ONE 13 2018 e0192450
Liu Y.J. Zhao L.H. Mosenthin R. Zhang J.Y. Ji C. Ma Q.G. Protective effect of vitamin E on laying performance, antioxidant capacity, and immunity in laying hens challenged with Salmonella Enteritidis Poult. Sci. 98 2019 5847 5854 31329983
Liu Y. Choe J. Kim S. Kim B. Campbell J.M. Polo J. Crenshaw J.D. Pettrigrew J.E. Song M. Dietary spray-dried plasma improves intestinal morphology of mated female mice under stress condition J. Anim. Sci. Tech. 60 2018 1 6
Livak K.J. Schmittgen T.D. Analysis of relative gene expression data using real-time quantitative PCR and the 2− ΔΔCT method Methd 25 2001 402 408
Miska K.B. Kahl S. Schreier L.L. Russell B. Kpodo K.R. Proszkowiec-Weglarz M. Delay of feed post-hatch causes changes in expression of immune-related genes and their correlation with components of gut microbiota, but does not affect protein expression Anims 12 2022 1316
National Research Council (NRC) Nutrient requirements of poultry: Ninth revised edition, 1994 1994 The National Academies Press Washington DC
Orimaye O.E. Ekunseitan D.A. Omaliko P.C. P.Fasina Y.O. Mitigation potential of herbal extracts and constituent bioactive compounds on salmonella in meat-type poultry Anim 14 2024 1087
Owusu-Asiedu A. Baidoo S.K. Nyachoti C.M. Marquardt R.R. Response of early-weaned pigs to spray-dried porcine or animal plasma-based diets supplemented with egg-yolk antibodies against enterotoxigenic Escherichia coli J. Anim. Sci. 80 2002 2895 2903 12462257
Pérez-Bosque A. Miró L. Amat C. Polo J. Moretó M. The anti-inflammatory effect of spray-dried plasma is mediated by a reduction in mucosal lymphocyte activation and infiltration in a mouse model of intestinal inflammation Nutri 8 2016 657
Pérez-Bosque A. Miró L. Polo J. Russell L. Campbell J. Weaver E. Crenshaw J. Moretó M. Dietary plasma proteins modulate the immune response of diffuse gut-associated lymphoid tissue in rats challenged with Staphylococcus aureus enterotoxin B J. Nutri. 138 2008 533 537
Pérez-Bosque A. Miró L. Polo L.J. Russell L. Campbell J. Weaver E. Crenshaw J. Moretó M. Dietary plasma protein supplements prevent the release of mucosal proinflammatory mediators in intestinal inflammation in rats J. Nutri. 140 2010 25 30
Petersen H. Nielsen J. Heegaard P.M.H. Application of acute phase protein measurements in veterinary clinical chemistry Vet. Res. 35 2004 163 187 15099494
Pierce J.L. Cromwell G.L. Lindemann M.D. Russell L.E. Weaver E.M. Effects of spray-dried animal plasma and immunoglobulins on performance of early weaned pigs J. Anim. Sci. 83 2005 2876 2885 16282627
Rodríguez S.P. Herrera A.L. Parra J.E. Gene expression of pro-inflammatory (IL-8, IL-18, TNF-α, and IFN-γ) and anti-inflammatory (IL-10) cytokines in the duodenum of broiler chickens exposed to lipopolysaccharides from Escherichia coli and Bacillus subtilis Vet. Wrld. 16 2023 564 570
Ruckman L.A. Petry A.L. Gould S.A. Patience J.F. The impact of porcine spray-dried plasma protein and dried egg protein harvested from hyper-immunized hens, provided in the presence or absence of subtherapeutic levels of antibiotics in the feed, on growth and indicators of intestinal function and physiology of nursery pigs Transl. Anim. Sci. 4 2020 1 16 32704961
Shaji S. Selvaraj R.K. Shanmugasundaram R. Salmonella infection in poultry: A review on the pathogen and control strategies Micro. 11 2023 2814
Shittu A. Raji A.A. Madugu S.A. Hassan A.W. Fasina F.O. Predictors of death and production performance of layer chickens in opened and sealed pens in a tropical savannah environment BMC Vet. Res. 10 2014 1 11 24383544
Swaggerty C.L. Kogut M.H. He H. Genovese K.J. Johnson C. Arsenault R.J. Differential levels of cecal colonization by salmonella enteritidis in chickens triggers distinct immune kinome profiles. front Vet. Sci. 4 2017 214
Touchette K.J. Carroll J.A. Allee G.L. Matteri R.L. Dyer C.J. Beausang L.A. Zannelli M.E. Effect of spray-dried plasma and lipopolysaccharide exposure on weaned pigs: I. Effects on the immune axis of weaned pigs J. Anim. Sci. 80 2002 494 501 11881933
Ulfman L.H. Leusen J.H. Savelkoul H.F. Warner J.O. Van Neerven R.J. Effects of bovine immunoglobulins on immune function, allergy, and infection Front. Nutri. 5 2018 52
Walters H.G. Jasek A. Campbell J.M. Coufal C. Lee J.T. Evaluation of spray-dried plasma in broiler diets with or without bacitracin methylene disalicylate J. Appl. Poult. Res. 28 2019 364 373
Wataha K. Menge T. Deng X. Shah A. Bode A. Holcomb J.B. Potter D. Kozar R. Spinella P.C. Pati S. Spray-dried plasma and fresh frozen plasma modulate permeability and inflammation in vitro in vascular endothelial cells Transfusion 53 2013 80S 90S 23301978
Wernicki A. Nowaczek A. Urban-Chmiel R. Bacteriophage therapy to combat bacterial infections in Poultry Viro. J. 14 2017 1 13
Zhang Y. Chen D.W. Yu B. He J. Yu J. Mao X.B. Wang J.X. Luo J.Q. Huang Z.Q. Zheng G.X. Zheng P. Spray-dried chicken plasma improves intestinal digestive function and regulates intestinal selected microflora in weaning piglets J. Anim. Sci. 93 2015 2967 2976 26115283
