Inducing inflammation in response to pathogen infection is known to be an energy-intensive process. An extended state of inflammation in production animals can be detrimental to performance parameters. Here, we compare two doses of a microencapsulated thymol-based feed additive blend and two different antibiotics in the context of a Salmonella Enterica subsp. Enterica serovar Enteritidis (SE) challenge. A total of 500, day-old, straight run chicks (Ross 708) were placed in floor pens (50 chicks/pen) and assigned to 5 groups with 2 replicates each. The groups were all fed with basal diets, without supplementation (Control) or supplemented with: tylosin at 25 g/MT (Tylosin); neomycin sulfate at 100 g/MT (Neomycin); or a microencapsulated thymol-based blend of botanicals at inclusions of 500 g/MT (Blend 500) or 1000 g/MT (Blend 1000). All the pens were orally challenged at day 4 with SE at 105 CFU/bird. At 7, 14, 21, and 35-days post hatch, ten animals from each pen were weighed and euthanized in order to culture enumerate SE in the ceca and determine SE prevalence in the liver. Gene based prevalence, gene expression, and kinome analysis were performed on jejunum samples collected at day 35. The higher dose of the tested botanical, Blend 1000, showed a statistically significant increase in final body weight relative to the other groups, indicating an energy benefit for this group. The Blend 1000 also showed a reduction in protein phosphorylation that corresponds to reduced inflammatory status that was unique compared to Blend 500 and the antibiotics. These birds also showed a clearance of orally inoculated SE demonstrating that reduced inflammation can benefit the broiler chicken in clearance of bacteria while maintaining growth.

Addition of vitamins and antioxidants has been long associated with increased immunity and are commonly used in the poultry industry; however, less is known regarding their use in broiler breeder hens. The objective of this study was to determine if feeding a complex of protected biofactors and antioxidants composed of vitamins and fermentation extracts to broiler breeder hens conferred resistance against Salmonella enterica serovar Enteritidis (S. Enteritidis) in the progeny chicks. Three-day-old chicks from control- and supplement-fed hens were challenged with S. Enteritidis and necropsied 4- and 11-days postchallenge (dpc) to determine if there were differences in invasion and colonization. Serum and jejunum were evaluated for various cytokine and chemokine production. Fewer (P = 0.002) chicks from supplement-fed hens had detectable S. Enteritidis in the ceca (32.6%) compared to chicks from control-fed hens (64%). By 11 dpc, significantly (P < 0.001) fewer chicks from supplement-fed hens were positive for S. Enteritidis (liver [36%]; ceca [16%]) compared to chicks from the control hens (liver [76%]; ceca [76%]). The recoverable S. Enteritidis in the cecal content was also lower (P = 0.01) at 11 dpc. In additional to the differences in invasion and colonization, cytokine and chemokine production were distinct between the 2 groups of chicks. Chicks from supplement-fed hens had increased production of IL-16, IL-6, MIP-3α, and RANTES in the jejunum while IL-16 and MIP-1β were higher in the serum of chicks from the control-fed hens. By 11 dpc, production of IFN-γ was decreased in the jejunum of chicks from supplement-fed hens. Collectively, these data demonstrate adding a protected complex of biofactors and antioxidants to the diet of broiler breeder hens offers a measure of transgenerational protection to the progeny against S. Enteritidis infection and reduces colonization that is mediated, in part, by a robust and distinct cytokine and chemokine response locally at the intestine and systemically in the blood.

Using a previously characterized and described abdominal model to define the avian immune response to Salmonella intra-abdominal challenge in chickens, we have adapted this technique for the study of chickens' immune response to a Campylobacter intra-abdominal challenge. The intra-abdominal Campylobacter infection model facilitates the characterization of peripheral blood leukocyte dynamics and abdominal cell infiltrates. Day-of-hatch Leghorn chickens were injected intra-abdominally (IA) with Campylobacter jejuni [(CJ)1 × 108 colony-forming units (CFUs)]. Changes in peripheral blood leukocyte numbers and abdominal cell infiltrates were monitored at 0, 4, 8, and 24 h post-injection. Peripheral blood leukocyte numbers were also determined for 2 h post-injection. For mortality studies, birds were injected intra-abdominally with 1 × 108 CFUs CJ and mortalities were recorded for 72 h post-injection. In the peripheral blood of CJ-injected chicks, total white blood cell (WBC) numbers began increasing by 2 h post-injection, peaking at 4 h post-injection with the predominant cell type being polymorphonuclear leukocytes (heterophils). Total WBCs declined after 8 h and this decline continued at 24 h, with total WBC numbers approaching control values. The injection of CJ into the abdominal cavity caused a rapid rise in abdominal cell infiltrates with the predominant infiltrating leukocytes being heterophils. Peak abdominal heterophil infiltrates were observed at 8 h post-injection, declining only slightly by 24 h post-injection. Mortality in the CJ challenge groups reached 37%. Mortality in the Salmonella enteritidis positive control groups were greater than 50%. The data suggest that Campylobacter infection does stimulate the innate immune response in chickens when administered IA, however, the immune response and infection is not characterized with the high levels of mortality observed with a Salmonella infection. These data provide a basis for a more definitive characterization of chickens' immune response to Campylobacter and a model to evaluate intervention strategies to prevent the infection and colonization of poultry.

Salmonella enterica is a group of facultative, gram-negative bacteria. Recently, new evidence indicated that Salmonella could reprogram the host metabolism to increase energy or metabolites available for intracellular replication. In this study, using a chicken-specific kinomic immunometabolism peptide array analysis, we found that infection by S. Enteritidis induced significant phosphorylation changes in many key proteins of the glycolytic pathway in chicken macrophage HD-11 cells, indicating a shift in glycolysis caused by Salmonella infection. Nitric oxide production and changes of glycolysis and mitochondrial oxidative phosphorylation (OXPHOS) represented by extracellular acidification rate (ECAR) and oxygen consumption rate (OCR), respectively, were measured in chicken macrophages infected with three Salmonella strains (S. Enteritidis, S. Heidelberg, and S. Senftenberg). The infection reduced glycolysis and enhanced OXPHOS in chicken macrophages as indicated by changes of ECAR and OCR. Salmonella strains differentially affected macrophage polarization and glycolysis. Among three strains tested, S. Enteritidis was most effective in downregulating glycolysis and promoting M2 polarization as measured by ECAR, ORC, and NO production; while S. Senftenberg did not alter glycolysis and may promote M1 polarization. Our results suggested that downregulation of host cell glycolysis and increase of M2 polarization of macrophages may contribute to increased intracellular survival of S. Enteritidis.

Poultry is a major source of human foodborne illness caused by broad host range Salmonella serovars (paratyphoid), and developing cost-effective, pre-harvest interventions to reduce these pathogens would be valuable to the industry and consumer. Host responses to infectious agents are often regulated through phosphorylation. However, proteomic mechanisms of Salmonella acute infection biology and host responses to the bacteria have been limited concentrating predominately on the genomic responses of the host to infection. Our recent development of chicken-specific peptide arrays for kinome analysis of host phosphorylation-based cellular signaling responses provided us with the opportunity to develop a more detailed understanding of the early (4-24 h post-infection) host-pathogen interactions during the initial colonization of the cecum by Salmonella. Using the chicken-specific kinomic immune peptide array, biological pathway analysis showed infection with S. Enteritidis increased signaling related to the innate immune response, relative to the non-infected control ceca. Notably, the acute innate immune signaling pathways were characterized by increased peptide phosphorylation (activation) of the Toll-like receptor and NOD-like receptor signaling pathways, the activation of the chemokine signaling pathway, and the activation of the apoptosis signaling pathways. In addition, Salmonella infection induced a dramatic alteration in the phosphorylation events of the JAK-STAT signaling pathway. Lastly, there is also significant activation of the T cell receptor signaling pathway demonstrating the initiation of the acquired immune response to Salmonella infection. Based on the individual phosphorylation events altered by the early Salmonella infection of the cecum, certain conclusions can be drawn: (1) Salmonella was recognized by both TLR and NOD receptors that initiated the innate immune response; (2) activation of the PPRs induced the production of chemokines CXCLi2 (IL-8) and cytokines IL-2, IL-6, IFN-α, and IFN-γ; (3) Salmonella infection targeted the JAK-STAT pathway as a means of evading the host response by targeting the dephosphorylation of JAK1 and TYK2 and STAT1,2,3,4, and 6; (4) apoptosis appears to be a host defense mechanism where the infection with Salmonella induced both the intrinsic and extrinsic apoptotic pathways; and (5) the T cell receptor signaling pathway activates the AP-1 and NF-κB transcription factor cascades, but not NFAT.

Necrotic enteritis (NE) is a devastating disease that has seen a resurgence of cases following the removal of antibiotics from feed resulting in financial loss and significant animal health concerns across the poultry industry. The objective was to evaluate the efficacy of a microencapsulated blend of organic (25% citric and 16.7% sorbic) acids and botanicals (1.7% thymol and 1% vanillin [AviPlusP]) to reduce clinical NE and determine the signaling pathways associated with any changes. Day of-hatch by-product broiler breeder chicks were randomly assigned to a control (0) or supplemented (500 g/MT) diet (n = 23-26) and evaluated in a NE challenge model (n = 3). Birds were administered 2X cocci vaccine on d 14 and challenged with a cocktail of Clostridium perfringens strains (107) on d 17 to 19. On d 20 to 21 birds were weighed, euthanized, and scored for NE lesions. Jejunal tissue was collected for kinome analysis using an immunometabolism peptide array (n = 5; 15/treatment) to compare tissue from supplement-fed birds to controls. Mortality and weight were analyzed using Student's t test and lesion scores analyzed using F-test two-sample for variances (P < 0.05). The kinome data was analyzed using PIIKA2 peptide array analysis software and fold-change between control and treated groups determined. Mortality in the supplemented group was 47.4% and 70.7% in controls (P = 0.004). Lesions scores were lower (P = 0.006) in supplemented birds (2.47) compared to controls (3.3). Supplement-fed birds tended (P = 0.19) to be heavier (848.6 g) than controls (796.2 g). Kinome analysis showed T cell receptor, TNF and NF-kB signaling pathways contributed to the improvements seen in the supplement-fed birds. The following peptides were signifi-cant (P < 0.05) in all 3 pathways: CHUK, MAP3K14, MAP3K7, and NFKB1 indicating their importance. Additionally, there were changes to IL6, IL10, and IFN-g mRNA expression in tissue between control-and supplement-fed chickens. In conclusion, the addition of a micro encapsulated blend of organic acids and botanicals to a broiler diet reduced the clinical signs of NE that was mediated by specific immune-related pathways.

Clostridium perfringens (Cp) is a Gram-positive anaerobe that is one of the causative agents of necrotic enteritis (NE) in chickens, which leads to high mortality. Owing to the ban of administering antibiotics in feed to chickens, there has been an increase in the number of NE outbreaks all over the world, and the estimated loss is approximately 6 billion U.S. dollars. The best alternative method to control NE without antibiotics could be vaccination. In this study, we exposed three different strains of Cp to electron beam (eBeam) irradiation to inactivate them and then used them as a killed vaccine to control the colonization of Cp in broiler chickens. The vaccine was delivered to 18-day old embryos in ovo and the chickens were challenged with the respective vaccine strain at two different time points (early and late) to test the protective efficacy of the vaccine. The results indicate that an effective eBeam dose of 10 kGy inactivated all three strains of Cp, did not affect the cell membrane or epitopes, induced significant levels of IgY in the vaccinated birds, and further reduced the colonization of Cp strains significantly (p < 0.0001) in late challenge (JGS4064: 4 out of 10; JGS1473: 0 out of 10; JGS4104: 3 out of 10). Further studies are necessary to enhance the efficacy of the vaccine and to understand the mechanism of vaccine protection.

The gut of food-producing animals is a reservoir for foodborne pathogens. Thymol is bactericidal against foodborne pathogens but rapid absorption of thymol from the proximal gut precludes the delivery of effective concentrations to the lower gut where pathogens mainly colonize. Thymol-β-d-glucopyranoside is reported to be more resistant to absorption than thymol in everted jejunal segments and could potentially function as a prebiotic by resisting degradation and absorption in the proximal gut but being hydrolysable by microbial β-glycosidase in the distal gut. Previous in vitro studies showed bactericidal effects of thymol-β-d-glucopyranoside against Campylobacter, Escherichia coli, and Salmonella enterica serovar Typhimurium in the presence but not absence of intestinal microbes expressing β-glycosidase activity, indicating that hydrolysis was required to obtain antimicrobial activity. Presently, the oral administration of thymol-β-d-glucopyranoside was studied to examine the effects on intestinal carriage of Campylobacter, E. coli, and S. Typhimurium in swine. The effects of thymol-β-d-glucopyranoside or thymol on antimicrobial sensitivity of representative E. coli isolates and characterized Salmonella strains were also explored. Results from two in vivo studies revealed little antimicrobial effects of thymol-β-d-glucopyranoside on Campylobacter, E. coli, or S. Typhimurium in swine gut. These findings add credence to current thinking that hydrolysis and absorption of thymol-β-d-glucopyranoside and thymol may be sufficiently rapid within the proximal gut to preclude delivery to the distal gut. Antibiotic susceptibilities of selected bacterial isolates and strains were mainly unaffected by thymol. Further research is warranted to overcome obstacles, preventing the delivery of efficacious amounts of thymol-β-d-glucopyranoside to the lower gut.

Strategies are sought to reduce the carriage and dissemination of zoonotic pathogens and antimicrobial-resistant microbes within food-producing animals and their production environment. Thymol (an essential oil) is a potent bactericide in vitro but in vivo efficacy has been inconsistent, largely due to its lipophilicity and absorption, which limits its passage and subsequent availability in the distal gastrointestinal tract. Conjugation of thymol to glucose to form thymol-β-d-glucopyranoside can decrease its absorption, but in vivo passage of effective concentrations to the lower gut remains suboptimal. Considering that contemporary swine diets often contain 5% or more added fat (to increase caloric density and reduce dustiness), we hypothesized that there may be sufficient residual fat in the distal intestinal tract to sequester free or conjugated thymol, thereby limiting the availability and subsequent effectiveness of this biocide. In support of this hypothesis, the anti-Salmonella Typhimurium effects of 6 mM free or conjugated thymol, expressed as log10-fold reductions of colony-forming units (CFU) ml−1, were diminished 90 and 58%, respectively, following 24-h in vitro anaerobic fecal incubation (at 39°C) with 3% added vegetable oil compared to reductions achieved during culture without added oil (6.1 log10 CFU ml−1). The antagonistic effect of vegetable oil and the bactericidal effect of free and conjugated thymol against Escherichia coli K88 tested similarly were diminished 86 and 84%, respectively, compared to reductions achieved in cultures incubated without added vegetable oil (5.7 log10 CFU ml−1). Inclusion of taurine (8 mg/ml), bile acids (0.6 mg/ml), or emulsifiers such as polyoxyethylene-40 stearate (0.2%), Tween 20, or Tween 80 (each at 1%) in the in vitro incubations had little effect on vegetable oil-caused inhibition of free or conjugated thymol. Based on these results, it seems reasonable to suspect that undigested lipid in the distal gut may limit the effectiveness of free or conjugated thymol. Accordingly, additional research is warranted to learn how to overcome obstacles diminishing bactericidal activity of free and conjugated thymol in the lower gastrointestinal tract of food-producing animals.

During the first week after hatch, young chicks are vulnerable to pathogens as the immune system is not fully developed. The objectives of this study were to determine if supplementing the starter diet with a microencapsulated feed additive containing citric and sorbic acids, thymol, and vanillin affects in vitro functional activity of peripheral blood leukocytes (PBLs). Day-old chicks (n = 800) were assigned to either a control diet (0 g/metric ton [MT]) or a diet supplemented with 500 g/MT of the microencapsulated additive. At 4 D of age, peripheral blood was collected (100 birds per treatment), and heterophils and monocytes isolated (n = 4). Heterophils were assayed for the ability to undergo degranulation and production of an oxidative burst response while nitric oxide production was measured in monocytes. Select cytokine and chemokine mRNA expression levels were also determined. Statistical analysis was performed using Student t test comparing the supplemented diet to the control (P ≤ 0.05). Heterophils isolated from chicks fed the microencapsulated citric and sorbic acids, thymol, and vanillin had higher (P ≤ 0.05) levels of degranulation and oxidative burst responses than those isolated from chicks on the control diet. Heterophils from the supplemented chicks also had greater (P ≤ 0.05) expression of IL10, IL1β, and CXCL8 mRNA than those from control-fed chicks. Similarly, nitric oxide production was significantly (P ≤ 0.05) higher in monocytes isolated from birds fed the supplement. The cytokine and chemokine profile in monocytes from the supplement-fed chicks showed a significant (P ≤ 0.05) drop in IL10 mRNA expression while IL1β, IL4, and CXCL8 were unchanged. In conclusion, 4 D of supplementation with a microencapsulated blend made up of citric and sorbic acids, thymol, and vanillin enhanced the in vitro PBL functions of degranulation, oxidative burst, and nitric oxide production compared with the control diet. Collectively, the data suggest feeding broiler chicks a diet supplemented with a microencapsulated blend of citric and sorbic acids, thymol, and vanillin may prime key immune cells making them more functionally efficient and acts as an immune-modulator to boost the inefficient and undeveloped immune system of young chicks.

Numerous Salmonella enterica serovars can cause disease and contamination of animal-produced foods. Oligosaccharide-rich products capable of blocking pathogen adherence to intestinal mucosa are attractive alternatives to antibiotics as these have potential to prevent enteric infections. Presently, a wood-derived prebiotic composed mainly of glucose-galactose-mannose-xylose oligomers was found to inhibit mannose-sensitive binding of select Salmonella Typhimurium and Escherichia coli strains when reacted with Saccharomyces boulardii. Tests for the ability of the prebiotic to prevent binding of a green fluorescent protein (GFP)-labeled S. Typhimurium to intestinal porcine epithelial cells (IPEC-J2) cultured in vitro revealed that prebiotic-exposed GFP-labeled S. Typhimurium bound > 30% fewer individual IPEC-J2 cells than did GFP-labeled S. Typhimurium having no prebiotic exposure. Quantitatively, 90% fewer prebiotic-exposed GFP-labeled S. Typhimurium cells were bound per individual IPEC-J2 cell compared to non-prebiotic exposed GFP-labeled S. Typhimurium. Comparison of invasiveness of S. Typhimurium DT104 against IPEC-J2 cells revealed greater than a 90% decrease in intracellular recovery of prebiotic-exposed S. Typhimurium DT104 compared to non-exposed controls (averaging 4.4 ± 0.2 log10 CFU/well). These results suggest compounds within the wood-derived prebiotic bound to E. coli and S. Typhimurium-produced adhesions and in the case of S. Typhimurium, this adhesion-binding activity inhibited the binding and invasion of IPEC-J2 cells.

With the reemergence of poultry diseases such as necrotic enteritis following the restriction of in-feed antibiotics, the search for antibiotic alternatives has become critically important. Postbiotics are non-viable bacterial products or metabolic byproducts from probiotic microorganisms that have positive effects on the host or microbiota. These are a promising alternative to antibiotics. Here, we describe the mechanism of action of a postbiotic in the context of a Clostridium perfringens (C. perfringens) challenge model. By using performance measurements and a peptide array kinome analysis, we describe the kinotypes and signal transduction changes elicited by the postbiotic with and without C. perfringens challenge. The postbiotic improves lesion scores, C. perfringens counts and mortality compared to challenge groups without the postbiotic, and it improves weight gain in the most severely challenged birds. The postbiotic predominantly affects the innate immune response and appears immunomodulatory. In the context of infection, it reduces the proinflammatory responses and generates a homeostatic-like response. This postbiotic is a viable alternative to antibiotics to improve poultry health in the context of C. perfringens pathogen challenge.

Strategies are sought to reduce the carriage and dissemination of zoonotic pathogens and antimicrobial resistant microbes within food-producing animals and their production environment. Thymol is an essential oil shown to be a potent bactericide in vitro but demonstration of its efficacy when fed to animals has been inconsistent, due largely to its lipophilicity which limits its passage and subsequent availability in the distal gastrointestinal tract. Conjugation of thymol to glucose to form thymol-β-D-glucopyranoside can decrease absorption of the conjugate, thereby promoting passage to more distal intestinal sites where pathogens primarily reside, yet in vivo efficacy of the conjugate remains suboptimal. It is possible that hydrolysis and absorption of thymol-β-D-glucopyranoside and free thymol may still have been rapid enough within the proximal small intestine to preclude their delivery to the cecum and large intestine. Considering that modern swine diets often contain 5 % or more fat, we hypothesized that even at 60 to 80 % apparent digestibility there may be passage of enough residual undigested lipid to the distal intestinal tract to sequester free or conjugated thymol within lipidic microenvironments, thereby limiting the availability and subsequent effectiveness of these biocides.

Calcium (Ca2+) is a pivotal intracellular second messenger and calmodulin (CaM) acts as a multifunctional Ca2+-binding protein that regulates downstream Ca2+ dependent signaling. Together they play an important role in regulating various cellular functions, including gene expression, maturation of phagolysosome, apoptosis, and immune response. Intracellular Ca2+ has been shown to play a critical role in Toll-like receptor-mediated immune response to microbial agonists in the HD11 chicken macrophage cell line. The role of that the Ca2+/CaM pathway plays in the intracellular survival of Salmonella in chicken macrophages has not been reported. In this study, kinome peptide array analysis indicated that the Ca2+/CaM pathway was significantly activated when chicken macrophage HD11 cells were infected with S. Enteritidis or S. Heidelberg. Further study demonstrated that treating cells with a pharmaceutical CaM inhibitor W-7, which disrupts the formation of Ca2+/CaM, significantly inhibited macrophages to produce nitric oxide and weaken the control of intracellular Salmonella replication. These results strongly indicate that CaM plays an important role in the innate immune response of chicken macrophages and that the Ca2+/CaM mediated signaling pathway is critically involved in the host cell response to Salmonella infection.

Salmonella Heidelberg is one of the most prevalent poultry Salmonella strains. Infection with S. Heidelberg strongly stimulates nitric oxide (NO) response in the chicken macrophage HD11 cells. In the present study, regulation of NO response to S. Heidelberg infection by the host mitogen-activated protein (MAP) kinases (p38, ERK1/2, and JNK) was investigated using the selective pharmaceutical inhibitors, SB203580, PO98059, and SP600125, respectively. At the concentrations used, the p38 inhibitor strongly inhibited the NO productions in S. Heidelberg infected cells (61-80% reduction; p≤0.05); whereas ERK1/2 inhibitor produced a much less inhibition (14-20% reduction; p≤0.05) and JNK inhibitor showed no inhibitory effect (4-6% increase). The total MAP kinases p38 and JNK were not changed by Salmonella infection, with mild increase detected in ERK1/2. However, large changes were observed in phopho-p38 (100% increase) and JNK (45% increase), with a minor change detected in phospho-ERK1/2 (9% increase). Although both p38 and JNK phosphorylation can be significantly inhibited (p≤ 0.05) by their inhibitors, only inhibition of p38 produced a significant reduction of NO. Together, these results indicated that p38 MAP kinase, not JNK and ERK1/2, plays a significant role in regulating the NO response of HD11 cells to Salmonella infection.

The intestinal tract harbors a diverse community of microbes that have co-evolved with the host immune system. Although many of these microbes execute functions that are critical for host physiology, the host immune system must control the microbial community so that the dynamics of this interdependent relationship is maintained. To facilitate host homeostasis, the immune system ensures that the microbial load is tolerated, but anatomically contained, while remaining reactive to microbial invasion. Inflammation is the most prevalent manifestation of host defense in reaction to alterations in tissue homeostasis and is elicited by innate immune receptors that recognize and detect infection, host damage, and danger signaling molecules that activate a highly regulated network of immunological and physiological events for the purpose of maintaining homeostasis and restoring functionality. The efficacy, duration, and consequences of an inflammatory response is dependent upon the type of trigger that is recognized by the innate immune receptors. Further, because of different triggers, there are multiple phenotypes of inflammation. Physiological inflammation is the homeostatic balance between tolerance of the microbiota and the reactivity to pathogen invasion. Pathologic inflammation is usually an acute response that involves the host response to toxins and infection often resulting in collateral damage to surrounding tissue and increased metabolic energy use. Metabolic inflammation is a chronic low-grade inflammation generated by excessive nutrient intake and the metabolic surplus fosters metabolic dysfunction by integrating signals from both the immune and metabolic systems. Sterile inflammation is a low-grade chronic inflammation, in the absence of an infection, in response to chemical, physical, and metabolic stimuli. With a sterile inflammatory response, the stimulus persists without being eliminated suggesting that collateral damage is the cause of the disease. The common denominator with all intestinal inflammatory phenotypes is the central role of the gut microbiota whether it be microbial balance and diversity of microbial metabolic production or microbial turnover.

EDITORIAL article Front. Vet. Sci., 20 June 2018Sec. Veterinary Infectious Diseases Volume 5 - 2018 | https://doi.org/10.3389/fvets.2018.00139

Protein kinases act in coordination with phosphatases to control protein phosphorylation and regulate signaling pathways and cellular processes involved in nearly every functions of cell life. Salmonella are known to manipulate the host kinase network to gain entrance and survive inside host cells. The effect of Salmonella infection on the host kinase network has been studied in mammalian cells, but information is largely lacking in chicken immune cells. Our previous study indicated that chicken macrophage cells respond differentially to different Salmonella strains. In order to better understand the interaction between chicken macrophages and Salmonella, we used a peptide array-based kinome analysis to identify cellular process and signaling pathways that may play a critical role in the outcome of Salmonella infection. The kinome assay was performed on chicken HD11 macrophages collected at 1.5, 3, and 7 h post-infection (hpi) with either S. Heidelberg or S. Enteritidis. A large number of peptides show significantly changed phosphorylation (p ≤ 0.05) during the infection: 390, 449, and 575 peptides for S. Enteritidis and 185, 470, and 442 for S. Heidelberg at 1.5, 3, and 7 hpi, respectively. Many pathways involved in immunity, signal transduction, cellular process, and metabolism were significantly altered, in some case differentially, during the infection by the two Salmonella strains. Particularly, effects on lysosome process, iNOS, CARD9, NLRP3, and MAPK pathway provide significant insight to the inter play between pathogens and chicken macrophage cells during the infection.

BACKGROUND Nigella sativa L. (NS) is a plant containing bioactive constituents such as thymoquinone. Extracts of NS improve performance and reduce enteropathogen colonization in poultry and small ruminants, but studies with swine are lacking. In two different studies oral administration of NS extracts at doses equivalent to 0, 1.5 and 4.5 g kg-1 diet was assessed on piglet performance and intestinal carriage of wildtype Escherichia coli and Campylobacter, and Salmonella Typhimurium. RESULTS Wildtype E. coli populations in the jejunal and rectal content collected 9 days after treatment began were decreased (P ≤ 0.05). Populations recovered from pigs treated with extract at 1.5 and 4.5 g kg-1 diet were 0.72-1.31 log10 units lower than the controls (ranging from 6.05 to 6.61 log10 CFU g-1 ). Wildtype Campylobacter and Salmonella Typhimurium were unaffected by NS treatment. Feed efficiency over the 9 days improved linearly (P < 0.05) from 3.88 with 0 NS-treated pigs to 1.47 and 1.41 with pigs treated with NS at 1.5 and 4.5 g kg-1 diet, respectively, possibly due to high glutamine/glutamic acid content of the NS extract. CONCLUSION NS supplementation of weanling pigs improved feed efficiency and helped control intestinal E. coli during this vulnerable production phase. © 2017 Society of Chemical Industry.

Campylobacter spp. are the second leading cause of bacterial-induced foodborne illnesses with an estimated economic burden of nearly $2B USD per year. Most human illness associated with campylobacteriosis is due to infection by C. jejuni and chickens are recognized as a reservoir that could lead to foodborne illness in humans resulting from handling or consuming raw or undercooked chicken. We recently developed a novel breeding strategy based on identification and selection of chickens with an inherently high and low phenotype of pro-inflammatory mediators including IL-6, CXCLi2, and CCLi2, hereafter referred to as the high and low lines, respectively. We have shown the high line chickens are more resistant to the foodborne and poultry pathogens Salmonella enterica serovar Enteritidis, Eimeria tenella, and Clostridium perfringens-induced necrotic enteritis compared to the low line. The objective of this study was to determine whether the same trend of enhanced resistance in the high line birds was observed for C. jejuni. Birds were challenged at 2 d of age by oral gavage (0.5 mL) with 5 × 106 colony forming units (cfu) of C. jejuni/mL, necropsied 4 d post challenge, and cecal content collected to determine if there was a difference in C. jejuni resistance between the high and low line chickens. There were fewer (P = 0.01) chickens from the high line (28/40 = 71.8%) that were colonized by C. jejuni compared to the low line (37/39 = 94.9%). The amount of C. jejuni recovered from the ceca of infected birds was quantified; however, no differences were observed (P = 0.10). Since the high line birds were also more resistant to C. jejuni, it provides additional validation of selection based on pro-inflammatory mediators producing a line of chickens with increased natural resistance against diverse foodborne and poultry pathogens. The poultry industry is moving towards reduced therapeutics and, as such, our breeding strategy would be a viable method to incorporate into traditional poultry breeding programs.