G. de Vries Reilingh

Kinetic and immunohistochemical characteristics of mitogen-induced cutaneous hypersensitivity in chickens selected for antibody responsiveness

Mitogen-induced cutaneous hypersensitivity was evaluated in chickens selected for high and low antibody responses to SRBC, and in a random bred control line. Wing web swelling responses were found after subcutaneous administration of phytohemagglutinin (PHA), concanavalin A (Con A), pokeweed mitogen (PWM), and Escherichia coli lipopolysaccharide (LPS), respectively, in all three lines. All mitogens induced significant acute 4 h wing web swelling responses, followed by a significant (classical) late 24 h wing web swelling response. The 4 h responses were significantly lower in the L line, whereas a tendency for lower responses at 24 h in the L line was found as well. Immunohistochemical evaluation of the early and late wing web swelling responses revealed extravascular localisation of leukocytes at 24 h after sensitization with mitogens, which consisted of CD4+ cells, CD8+ cells, TCR-1+ cells, and heterophils, but no B cells, whereas the 4 h swelling response was primarily characterized by oedema. Cutaneous hypersensitivity either initiated by T-cell mitogens as well as B-cell mitogens may depend for an important part on the rapid induction of local homing of lymphocytes towards the sensitizing agent, which may be mediated by an acute local expression of molecules with chemo-attractive capacities. Interpretation of cellular immunity responses in vivo such as delayed-type hypersensitivity should therefore incorporate oedema-initiating characteristics of sensitizing agents. The relationship between the magnitude of cutaneous hypersensitivity to mitogens and selection for antibody responsiveness is discussed.

Effects of genetic origin and social environment on behavioral response to manual restraint and monoamine functioning in laying hens.

Purebred laying hen lines of White Leghorn (WL) origin have been found to be more flighty and to show more feather pecking than lines of Rhode Island Red (RIR) origin. It has been found, however, that when RIR birds were housed together with WL birds, RIR birds became more flighty and those mixed groups developed more feather damage than pure-line cage-housed groups. It is unknown, however, whether this effect of social environment is accompanied by changes in stress-related behavior and neurophysiological activity, which are assumed to be associated with increased feather damage. The objective of this study was therefore to investigate the effects of genetic origin (WL or RIR) and social environment (mixed or pure groups) on behavioral response to manual restraint and monoamine functioning. Monoamine functioning was measured by brain serotonin (5-HT) and dopamine turnover. Furthermore, correlations between 5-HT turnover in the brain and peripheral measures of 5-HT in the blood were calculated. Experimental birds, housed either with other birds from the same genetic origin (pure groups) or with both RIR and WL birds (mixed groups) from hatching onward, were subjected to a manual restraint test at 47 wk of age. The WL birds struggled less during restraint and had higher dopamine and 5-HT turnover levels after restraint than did RIR birds. The WL birds also showed higher levels of platelet 5-HT uptake than did RIR birds. No effects of social environment were found. Blood and brain 5-HT measures were found to be correlated, with correlations ranging from 0.34 to 0.57, which seems to offer opportunities for less invasive peripheral indicators of 5-HT activity. In conclusion, genetic origin, but not social environment, affected the behavioral response to manual restraint and monoamine functioning in laying hens.

Immune responses of piglets to Pasteurella multocida toxin and toxoid

Experimental atrophic rhinitis (AR), serum antibody titres and in vitro lymphoproliferation to Pasteurella multocida derived toxin (Pm-T) were studied in piglets. Specific immune responses to Pm-T and Pm-T induced conchae atrophy were compared with AR immunity. This immunity was initiated by the Nobi-VAC AR-T vaccine administered at various times with respect to Pm-T challenge. Animals challenged with Pm-T developed conchae atrophy, but no antibodies nor cellular immune responses to Pm-T were detected. Vaccination 3 weeks before Pm-T challenge protected pigs against breakdown of nasal bony tissues. This protection was accompanied by an increase of serum antibodies and in vitro lymphoproliferation to Pm-T. Animals vaccinated 10 days before or after Pm-T challenge also had antibodies and cellular immune responses. However, these animals developed AR. In vitro, Pm-T appeared mitogenic for quiescent (non-immune) peripheral lymphocytes and Concanavalin A stimulated lymphocytes from some pigs. These in vitro lymphoproliferative responses could be partly abrogated by the addition of monomorphic anti-swine major histocompatibility complex class II DQ and DR specific monoclonal antibodies. We conclude that Pm-T is poorly immunogenic in vivo and does not initiate a protective Pm-T specific immune response. Pigs were protected from AR by vaccination, but protection was dependent on the timing of vaccine administration. We speculate that Pm-T modifies the immune response such that the response is not directed towards the toxin but to an unidentified component in the nose of piglets.

Natural antibodies related to metabolic and mammary health in dairy cows

Natural antibodies (NAb) are defined as antibodies that circulate in normal healthy individuals under the absence of deliberate antigenic stimulation. Two types of NAb are distinguished: NAb towards exogenous antigens and NAb towards autoantigens (N(A)Ab). The objectives of the current study were threefold. First, we studied the relation between metabolic health and concentrations of NAb binding keyhole limpet hemocyanin (KLH) or lipopolysaccharide (LPS) in milk and plasma of dairy cows in early lactation. Second, we determined the presence of N(A)Ab binding transferrin, myosin and thyroglobulin in bovine milk. Third, we studied the relation between N(A)Ab in bovine milk and mammary health. For the first objective, dairy cows were either fed a control (C) (n=8) or a diet where 2 kg of concentrates were replaced by an iso-energetic concentrate containing marine algae (ALG) from week -3 till 8 postpartum (experiment 1). Plasma and milk samples were analyzed weekly for NAb binding either KLH or LPS. Plasma was analyzed for glucose, non-esterified fatty acids (NEFA) and β-hydroxybutyrate (BHBA). For the second and third objective, milk samples were collected weekly from 96 dairy cows from week 2 till 9 postpartum and analyzed for milk composition and N(A)Ab binding myosin, transferrin and thyroglobulin (experiment 2). For both datasets, N(A)Ab titers are expressed as (2)log values of the highest dilution giving a positive reaction. Data are expressed as means ± SEM. Repeated observations were analyzed in a mixed model. In experiment 1, no diet effect (P>0.05) was observed on NAb binding LPS in plasma or milk, NAb binding KLH in milk was greater (P=0.05) for cows fed the control diet. Concentration of NAb binding KLH and LPS in plasma was negatively related to plasma NEFA concentration (P<0.05). In experiment 2, NAb binding myosin (5.66 ± 0.06), thyroglobulin (4.85 ± 0.06), and transferrin (5.76 ± 0.07) were identified in milk. Clinical mastitis incidence (9%) tended to be positively related to concentration of NAb binding myosin (P=0.06) and negatively related to Nab binding transferrin (P=0.08). In conclusion, NAb binding KLH and LPS in plasma and milk are related to metabolic health, as indicated by plasma NEFA concentration. Furthermore, this study demonstrates the presence of N(A)Ab in bovine milk and shows trends for a relation between N(A)Ab binding auto-antigens and mastitis. Future studies should confirm these trends and shed light on the predictive value of N(A)Ab in bovine milk for mammary health.

Does enhancement of specific immune responses predispose laying hens for feather pecking

To mimic airborne immune challenges, layer hens were intratracheally and concurrently challenged with various doses of the protein antigen human serum albumin (HuSA) and the pathogen-associated molecular pattern lipopolysaccharide (LPS) at 7 and 13 wk of age. All groups received 1 similar dose of HuSA plus LPS at 11 mo of age. Evaluation of plumage and body condition at 12 mo of age revealed that birds that had undergone intratracheal immunization with a high dosage of HuSA, irrespective of the concurrent dose of LPS, had significantly more feather damage but less wounds to the vent region, as opposed to birds not receiving HuSA. On the other hand, a high dosage of LPS was related to comb damage. These results suggest that stimulation of specific (humoral) immune responses (to HuSA) rather than innate responses (to LPS) at a young age may predispose layers for feather pecking (FP) behavior at later ages. Involvement of immune mechanisms in FP or vent damage may differ. Predisposal for FP behavior by specific immunity can have consequences for health and vaccine management.

Effect of early life thermal conditioning and immune challenge on thermotolerance and humoral immune competence in adult laying hens

Effects of early life experience with climatic (heat) and hygienic [lipopolysaccharide (LPS)] stress on adaptability to the same stressors in later life were studied in laying hens. Chicks were exposed to 37 degrees C for 24 h at d 5 of age (n = 12) or were i.v.-administered once with 1 mg/kg of BW of LPS at 6 wk of age (n = 12), whereas a control group was reared under standard conditions receiving a placebo treatment of PBS (n = 36). At 24 wk of age, hens treated in early life were reexposed to the same stressor. Early life control hens were exposed to heat stress (n = 12), i.v.-administered with LPS (n = 12), or not exposed (n = 12). To evaluate improvement of adaptability, effects of climatic and hygienic stress on performance, humoral immune competence, and endocrine responsiveness were investigated in hens with early life experience to the stressors and hens only exposed to the stressors in later life. Early life heat exposure did not affect performance, immune, and endocrine parameters. Treatment x time interactions were found for level of antibody (Ab) binding to LPS and keyhole limpet hemocyanin (KLH) after LPS administration, indicating that hens with early life LPS experience differed in response level (Ab binding to LPS) and response pattern (Ab binding to LPS and KLH) compared with hens administered with LPS only at adult age. Our data suggest that early life heat stress exposure did not affect adaptability of laying hens to heat stress in later life. However, early life LPS exposure affected kinetics and magnitude of Ab levels binding to LPS and KLH, indicating that early life LPS exposure can enhance the status of immune reactivity or induce a higher sensitivity to LPS.

Effect of three different housing systems on immune responses and body weight of chicken lines divergently selected for antibody responses to sheep red blood cells

Abstract There is an urgent need for knowledge on the effects of alternative housing conditions on parameters of health and production in current food animals, which are usually selected for a prolonged period for production traits. Thus, it is as yet unknown, whether genotype or environmental factors, or both, are major determinants of these traits. Therefore, as a model, the effects of keeping layers in three different types of housing: solitary in battery cages (BC), on a colony basis in a barn provided with litter and perches (BPL) and free range housing with access to the outdoors (FR), respectively, on immune competence and body weight (gain) of chicken lines divergently selected for high (H line) or low (L line) antibody responses to sheep red blood cells (SRBCs), next to a random bred control (C) line were studied. Antibody responses to keyhole limpet haemocyanin (KLH), Escherichia coli lipopolysaccharide (LPS) and ovalbumin (OVA) were significantly higher in the H line as compared to the C and L lines in all three housing systems, whereas in vitro mitogenesis of peripheral blood leukocytes to concanavalin (Con) A, and body weight (BW) were significantly higher in the L line in all three housing systems. No differences between the lines were found with respect to phagocyte activity. Housing conditions affected antibody responses to KLH, i.e. birds kept solitary in BC, and birds kept in BPL systems showed significantly higher antibody responses to KLH, than birds kept in the FR system. Also in vitro T-cell responses to KLH were significantly higher in BC birds than in the FR birds, but T cell responses to Con A were higher in the FR birds. Antibody responses to KLH, but also LPS and OVA were significantly affected by a housing×time interaction. Titers to these three antigens remained at a higher level in the BC then in the FR system. Birds kept in battery cages were heavier, and produced their first eggs approximately 2 and 3 weeks earlier than birds in the BPL and the FR system, respectively. The absence in the present study of significant interactions between line and housing systems on immune competence suggests that regardless of genotype, the animals respond similarly to different environments. The modulation of the magnitude of specific, but not innate immune responses in the three lines by environmental (housing) conditions suggest that the environment may favour types of specific immune responses. In addition, our data suggest that specific genotypes and specific environments may be additive in disease resistance.

Effect of aging and repeated intratracheal challenge on levels of cryptic and overt natural antibodies in poultry

Natural antibodies (NAb) have been divided in 2 classes: overt and cryptic. Overt NAb can be detected in unfractionated normal sera of nonimmunized mammals and chickens. Cryptic NAb as described in mammals need an in vitro physical or biochemical treatment to be detected, which may reflect their biochemical modification in situ during inflammation or infection. We studied the effect of concurrent primary, secondary, and tertiary intratracheal (i.t.) challenges with lipopolysaccharide (LPS) and human serum albumin on levels of 2 cryptic NAb [i.e., NAb binding actin (ACT), or thyroglobulin (THYRO)] and levels of an overt NAb binding keyhole limpet hemocyanin (KLH). In addition, effect of aging of the birds on levels of NAb was taken into account. Presence and changes on the level of the cryptic antibodies directed to ACT and THYRO were expected after the i.t. challenges with LPS (and human serum albumin). However, levels of NAb binding ACT were only significantly enhanced by LPS after primary challenge, whereas levels of NAb binding THYRO were not affected by primary nor secondary challenges but were significantly decreased by LPS after the third immunization. On the other hand, no changes in the levels of overt NAb binding KLH were expected, but levels of NAb binding KLH were significantly enhanced after the primary and secondary challenges with LPS. Levels of all three NAb increased with aging, but the different challenges performed at 3 moments during aging did not significantly or consistently affect levels of the overt nor cryptic NAb. Our results suggest that chickens might react by overt NAb as well as cryptic NAb to an infection or inflammation rather than by cryptic NAb only. The relation between various types of NAb and i.t. immunization with antigen and LPS and aging is discussed.

Long-term effects of early life microbiota disturbance on adaptive immunity in laying hens

Due to an interplay between intestinal microbiota and immune system, disruption of intestinal microbiota composition during immune development may have consequences for immune responses later in life. The present study investigated the effects of antibiotic treatment in the first weeks of life on the specific antibody response later in life in chickens. Layer chicks received an antibiotic cocktail consisting of vancomycin, neomycin, metronidazole, and amphotericin-B by oral gavage every 12 h, and ampicillin and colistin in drinking water for the first week of life. After the first week of life, chicks received ampicillin and colistin in drinking water for two more weeks. Control birds received no antibiotic cocktail and plain drinking water. Fecal microbiota composition was determined during antibiotic treatment (d 8 and 22), two weeks after cessation of antibiotic treatment (d 36), and at the end of the experimental period at d 175 using a 16S ribosomal RNA gene targeted microarray, the Chicken Intestinal Tract Chip (ChickChip). During antibiotic treatment fecal microbiota composition differed strongly between treatment groups. Fecal microbiota of antibiotic treated birds consisted mainly of Proteobacteria, and in particular E.coli, whereas fecal microbiota of control birds consisted mainly of Firmicutes, such as lactobacilli and clostridia. Two weeks after cessation of antibiotic treatment fecal microbiota composition of antibiotic treated birds had recovered and was similar to that of control birds. On d 105, 12 weeks after cessation of antibiotic treatment, chicks of both treatment groups received an intra-tracheal lipopolysaccharide (LPS)/human serum albumin (HuSA) challenge. Antibody titers against LPS and HuSA were measured 10 days after administration of the challenge. While T cell independent antibody titers (LPS) were not affected by antibiotic treatment, antibiotic treated birds showed lower T cell dependent antibody titers (HuSA) compared with control birds. In conclusion, intestinal microbial dysbiosis early in life may still have effects on the specific antibody response months after cessation of antibiotic treatment and despite an apparent recovery in microbiota composition.

Development of ileal cytokine and immunoglobulin expression levels in response to early feeding in broilers and layers

Provision of feed in the immediate posthatch period may influence interaction between intestinal microbiota and immune system, and consequently immunological development of the chick. This study addressed ileal immune development in response to early feeding in 2 chicken breeds selected for different production traits: broilers and layers. Chicks of both breeds either received feed and water immediately posthatch or were subjected to a 72-h feed and water delay. Ileal cytokine and immunoglobulin mRNA expression levels were determined at different time points. Effects of early feeding were limited, but breeds differed strikingly regarding cytokine and immunoglobulin expression levels. Cytokine expression levels in broilers were low compared with layers and showed a transient drop in the second to third week of life. In contrast, broilers showed considerably higher expression levels of IgA, IgM, and IgY. These findings indicate that the 2 breeds use different immune strategies, at least on the ileal level.