Immunological parameters in meat-type chicken lines divergently selected by antibody response to Escherichia coli vaccination.

Our group has established two lines of meat-type chickens divergently selected for early (HC line) and late (LC line) antibody responsiveness at 10 days of age to immunization with inactivated pathogenic Escherichia coli bacteria. The question addressed in the study presented here is whether this selection has changed other immunological responses, increasing the overall 'early' immunocompetence. Broilers of the third and fourth generations (S3 and S4) of the selected lines (HC and LC) and a control, unselected line (CT) were vaccinated at 10 days of age with E. coli vaccine, Newcastle virus vaccine (NDV), sheep erythrocytes (SRBC) or bovine serum albumin (BSA). Line-HC chicks exhibited higher antibody titers to E. coli, NDV and SRBC than CT or LC chicks. At 20 days of age HC chicks demonstrated a higher total protein and a higher beta- and gamma-globulin levels in their serum. At 21 days of age, HC chicks cleared carbon particles faster than LC chicks. Peripheral blood lymphocytes (PBL) from HC chicks vaccinated with E. coli vaccine, proliferated in vitro more actively in the presence of the stimulating antigen than the PBL of LC chicks. Peripheral blood lymphocytes (PBL) obtained from HC-line chicks exhibited a higher proliferative response to concanavalin A (Con A)-, phytohemagglutinin (PHA)- or pokeweed mitogen (PWM)-stimulation than LC PBL. These results demonstrate that the selection for high or low antibody response to E. coli at a young age resulted also in a significant change in the response of other parameters of the immune system. The high response to E. coli was found to be associated with a high antibody response to other antigens (NDV and SRBC), increased phagocytic activity and increased proliferative response to antigen or mitogens. The selection most probably affected early immunocompetence.     

Exploration of stress-induced immunosuppression in chickens reveals both stress-resistant and stress-susceptible antigen responses

In the present study, depriving chickens of foraging material was shown to induce stress. The impact of this type of stress on the immune response was compared with feeding of corticosterone (1.5 mg per bird per day), a hormone known to be immunosuppressive and to be the major stress hormone of chickens. Corticosterone feeding induced stress as revealed by higher heterophil/lymphocyte (H/L) ratios, longer tonic immobility (TI) reaction, reduced body weight gain and reduced egg production. Blood corticosterone levels were increased. Corticosterone feeding decreased the antibody response to tetanus toxoid and SRBC, DTH to PPD from Mycobacterium tuberculosis and the inflammatory response to PHA. Housing chickens on slats also induced chronic stress, as evidenced by increased H/L ratios, prolonged TI duration and decreased egg production. Corticosterone levels were slightly but not significantly enhanced. This novel form of chronic stress strongly suppressed humoral and cellular immune responses as evidenced by lower antibody titers to sheep red blood cells (SRBC) and tetanus toxoid (TT) decreased DTH reaction to PPD and inflammatory reaction to PHA in the skin. In contrast, the antibody response to human serum albumin (HSA) was neither influenced by corticosterone feeding nor by keeping the birds on slats. Even the combination of corticosterone feeding and housing the birds on slats did not significantly impair antibody responses to HSA. In conclusion, the present study showed that chronic stress induced by depriving the birds of foraging material led to a similar impairment of humoral and cell-mediated immunity as did feeding with corticosterone. More importantly, it showed for the first time that depending on the antigen tested, there are stress-resistant and stress-susceptible antigen responses.     

Immune mechanisms in Marek's disease.

Resistance to progressive tumor development in MD is either naturally inherited or can be induced by vaccination with apathogenic or attenuated MDV or with HVT. Studies on the effects of immunosuppression on resistance have shown that natural and vaccine induced resistance may be mediated through immune responses. Cell-mediated immune responses rather than humoral responses appear to be of principal importance. The antigen(s) against which protective cell-mediated immunity is elicited are not yet clearly delineated. Both virus-related and tumor antigens may be involved. Progress in the understanding of cell-mediated immunity in MD has been slow because of lack of reproducible in vitro tests to measure this response in infected chickens. The development of lymphoblastoid cell lines from MD lymphomas, however, has enabled the development of an in vitro cytotoxicity test. In this test, which utilizes MSB-1 cells as the target cells, a specific cell-mediated immune response, presumably against the tumor antigen, MATSA, was detected in chickens infected with MDV. Further studies using similar in vitro tests will facilitate a better understanding of the role cell-mediated immune responses might play in development of MD.     

Diminished antibody response to rinderpest vaccination in cattle undergoing experimental East Coast fever.

The immunocompetence of cattle undergoing East Coast Fever (ECF) reactions of varying degrees of intensity was evaluated using the neutralising antibody response of these cattle to the vaccine response was found in animals undergoing severe ECF reactions. The results suggest that the massive lymphoid cell involvement of such severe ECF reactions diminishes the immune responsiveness of cattle to rinderpest vaccine.     

Effects of direct immersion in antigen on immunological memory in young carp, Cyprinus carpio

The study asks whether, in fish, antigens encountered early in life can prime the immune system to yield memory responses on subsequent challenge with the same antigen and, if so, whether positive immunity or immunological tolerance is induced. The direct immersion method of vaccination was used to prime 4 week old carp, Cyprinus carpio, and was compared with priming by injection. Three different forms of antigen were used: the thymus dependent antigen, human gamma globulin (HGG) in soluble and in particulate (latex bound) form; also the putative thymus independent bacterin, formalin-killed Aeromonas salmonicida. The thymus dependent antigens were also used on 9 month old animals. In 4 week old carp, A. salmonicida vaccine delivered either by direct immersion or intraperitoneally (i.p.) yielded enhanced serum antibody levels and heightened proliferative responses in the lymphoid tissue of the spleen and kidney. Latex-bound HGG applied by direct immersion was found to partially suppress secondary antibody production while still eliciting enhanced proliferation. The decrease in antibody production following direct immersion priming of young fish with latex-bound HGG was not nearly as marked as the tolerance induced following priming with latex-bound HGG by the i.p. route and, unlike the tolerance induced by the injection route, may possibly still occur in older fish. When HGG was applied to young carp in soluble form by direct immersion it was ineffective and failed to influence memory induction. This is in contrast to the antibody tolerance, accompanied by an enhanced proliferative response following challenge, which resulted from administration of the soluble antigen by injection in the young fish. The status of the immune system in these antibody-tolerant fish is still far from clear. This highlights the need for further investigation of the role of cell-mediated reactions and local immunity in the immune responses of fish.     

The influence of T. evansi infection on the immuno-responsiveness of experimentally infected water buffaloes.

In order to define the immuno-suppressive capacity of Trypanosoma evansi infections in buffaloes on the induction of immune responses against heterologous antigens, infected and non-infected buffaloes were vaccinated against Pasteurella multocida (haemorrhagic septicemia) and were simultaneously immunised with a control antigen, human serum albumin (HSA). Antibody responses against HSA were significantly reduced in T. evansi-infected animals, but no conclusive data were obtained on the antibody responses against P. multocida. Conversely, the local inflammatory response at the site of Pasteurella vaccination, as measured by increase in size, was significantly reduced in T. evansi-infected animals.These results indicate that the inductive capacity to mount humoral and cell-mediated immune responses against heterologous antigens is suppressed in T. evansi-infected animals. Consequently, T. evansi infection might interfere with the development of protective immunity upon heterologous vaccinations and could explain the poor protection of Pasteurella-vaccinated buffaloes in T. evansi-endemic areas of Vietnam.     

Immunological differences between layer- and broiler-type chickens

In commercial poultry husbandry, alternatives for the use of antibiotics and vaccines are under investigation, which preferably have to be applicable for both layer- and broiler-type chickens. There are indications that the defense mechanisms vary between layer- and broiler-type chickens. Therefore, the difference in immune response between layer- and broiler-type chickens of the same age was investigated, using TNP-KLH (trinitrophenyl-conjugated keyhole limpet hemocyanin) as antigen without adjuvant. First different routes of immunization (intravenously, intramuscular, subcutaneous and ocular) were examined to find out which immunization route gives the highest antibody titers. The intravenous immunization route resulted in higher TNP-specific antibody responses than the other immunization routes tested and therefore this immunization route was used in both following experiments. In order to investigate the optimal dose of antigen needed for immunization, a dose-response curve in broiler- and layer-type chickens was completed. The humoral immune response was measured in serum by a TNP-specific ELISA and the in vitro cellular immune response by an antigen-specific lymphocyte proliferation assay.The antibody response of layer- and broiler-type chickens appeared to differ, not only in optimal dose and response, but also in kinetics of the response itself. Broiler chickens generated higher IgM anti-TNP titers whereas layer-type chickens generated higher IgG anti-TNP titers. This specific antibody response in broiler-type chickens did not last as long as in layer-type chickens. The TNP-specific cellular immune response was detectable in layer-type chickens, but not in broilers. Both types generate a non-specific cellular immune response, although this response in broilers is lower than in layer-type chickens.From these results, we conclude that broilers primarily respond to TNP-KLH with a high IgM antibody response whereas layer-type chickens respond with a high IgG response. In addition, the cellular response of layer-type chickens is much higher than the response of broilers. The results suggest that broilers are specialized in the production of a strong short-term humoral response and layer-type chickens in a long-term humoral response in combination with a strong cellular response, which is in conformity with their life expectancy.     

Nutrition--mechanisms of immunosuppression.

Nutritionists must formulate diets that supply adequate amounts of nutrients from five major groups. These are carbohydrate, protein (amino acids), fat, minerals and vitamins. Carbohydrate is usually a cheaper source of energy than fat, but fat is often used to increase the caloric concentration of the diet. Variations in energy intake which may effect immunocompetence usually result from management practices rather than diet formulation. Feed restriction for broiler breeders and withholding feed in forced molting practices may affect immunocompetence. Feed restriction causes higher plasma corticosterone levels, which are known to decrease the immune response, possibly through effects on cytokines. Excessive feed, through forced feeding, may also have short-term effects on indicators of humoral immunity. Protein and amino acid nutrition have been studied in relation to immunocompetence. The level of dietary amino acid needed to maximize growth and feed efficiency will also generally maximize measures of immunocompetence. The level of amino acids needed for maximum growth is lower in chicks which have been immunologically stressed than in chicks which have not. An immune response changes metabolism so that less growth occurs, thereby decreasing the need for amino acids. Dietary levels of minerals can affect immunocompetence. While deficient levels of sodium and chloride decrease humoral immunity, levels of these nutrients which supported maximum growth also supported maximal humoral immunity. Low dietary zinc levels did not affect indicators of immunocompetence in the chick. The effect of fat soluble vitamin levels on the immune system has been studied. Vitamin A is needed to maintain epithelial tissue and prevent infection. Cellular immune response is decreased when the chick is deficient in this nutrient. Several indicators of immune responsiveness are depressed when chicks are vitamin E and/or selenium deficient. Since these nutrients serve as antioxidants, cellular integrity may be affected by a deficiency. Cellular integrity is very important for receiving, and responding to the messages needed to coordinate an immune response. High levels of vitamin E (greater than 10 times the required level) have been found to be immunostimulatory.     

Protective immunity against infectious bursal disease virus in chickens in the absence of virus-specific antibodies

The role of cell-mediated immunity (CMI) in pathogenesis of infectious bursal disease virus (IBDV) was investigated. One-day-old specific pathogen-free chickens were treated with 3mg of cyclophosphamide (Cy) per chicken for 4 consecutive days and, 3 weeks later, infected with the IBDV-IM strain. Chickens were examined for: (a) mitogenic response of splenocytes to ConA, as an indicator of T-cell functions in vitro, (b) antibody against IBDV by ELISA, (c) IBDV genome in various tissues by RT-PCR and (d) immunological memory. At the time of IBDV infection, Cy-treated chickens had depleted bursal tissue (an avian primary B-cell lymphoid organ), severely compromised antibody-producing ability, but normal T-cell response to ConA. In primary infection, no detectable antibody against IBDV antigen in Cy-treated, IBDV-infected chickens was observed up to 28 days post-infection (PI), while IBDV genome was detected by RT-PCR in spleen, thymus, liver and blood until 10 days PI. Like intact control chickens infected with IBDV, Cy-treated, IBDV-infected chickens suppressed splenocytes responses to ConA from 5 to 10 days PI, suggesting that intact control as well as Cy-treated chickens responded similarly to IBDV infection in the early phase. Following re-infection with IBDV, no detectable secondary antibody response to IBDV as well as IBDV genome in tissues were observed in Cy-treated chickens, while intact control chickens developed vigorous secondary antibody response. Similar to intact control chickens infected with IBDV, Cy-treated chickens after second infection with IBDV did not suppress splenocyte response to ConA. These results suggested that in the absence of detectable anti-IBDV antibodies, protection of Cy-treated chickens from IBDV infection may occur via immunological memory mediated by CMI. We concluded that under normal conditions, IBDV induces a protective antibody response, however, in the absence of antibody, CMI alone is adequate in protecting birds against virulent IBDV.     

Immune modulation: the genetic approach

As part of a comprehensive strategy to combat diseases, improving genetically resistance to diseases and therefore immune capacities of animals is more and more desirable. However, research is still needed to develop genetic tools that may be used. In this search, lines selected for various immune responses are used to study relevant immune markers. Chickens have been selected for six generations for three different in vivo immune responses: high antibody response, high cell-mediated immune response and high phagocytic activity. Each line, selected for one trait, showed significant increase in immune capacity for this trait. In addition, results showed clearly independence between the three immune responses analyzed, meaning that a global approach is needed to improve immune capacity. Selected lines allow to follow genetic markers linked to immune response genes. In the different lines, different patterns in MHC gene frequency were observed and MHC alleles differed in their effects according to the immune trait. Some correlations were found between immune responses and production traits. The selected lines will be used to find other "known" immune response genes or "anonymous" genetic markers, which may become the future tools to modulate immune responsiveness of animals.     

T lymphocyte subpopulations diverge in commercially raised chickens

To evaluate immunocompetence in commercially raised chickens, we immunophenotyped Dekalb Delta and H&N White Leghorn (WLH) hybrids, 20 chickens in each of 3 age groups (9 wk [juvenile], 25 wk [young adult], and 79 or 80 wk [adult]), for circulating CD3+, CD4+, CD8+, TCR1+, TCR2+, and TCR3+ lymphocytes. The proportion of CD3+ T cells, including CD4+ and CD8+ subsets, was increased in the hybrids as compared with published values for laboratory-raised outbred WLH chickens. The proportion of the TCR2+ (Vbeta1) T cell subpopulation was also increased. An age-related decrease in the proportion of TCR1+ (gammasigma) T cells was noted in both hybrids. Further, a remarkably low CD4:CD8 ratio was evident in all age groups of both hybrids, indicating decreased immunocompetence. Overall, these experiments provide age-related proportions of various peripheral-blood T lymphocyte subpopulations in commercially raised Dekalb Delta and H&N chickens that diverge from the proportions in laboratory-raised outbred WLH chickens and suggest reduced immunocompetence. Such a decline in immunocompetence, including humoral immune capacity, could be attributed to genetic selection for production traits, environmental factors associated with commercial operations, and intense immunization.     

Comparison of natural resistance in seven genetic groups of meat-type chicken

1. Several studies have shown that genetic variation exists in response to various Salmonella strains in mammals and poultry. In the current study immunocompetence traits related to natural resistance to Salmonella were measured in 7 genetic groups of meat-type chickens (in total 296 chickens involved). 2. Variables were measured of both innate (phagocytic activity) and adaptive immune responses that are important after a natural or experimental Salmonella enteritidis infection. Two traditional Old Dutch Breeds (groups 1 and 2), four commercial broiler groups (groups 3 to 6), and one experimental broiler group (group 7) were used. In two periods, birds of each group were killed for examination at ages between 14 and 35 d post hatch. 3. Significant differences between groups were found for most immune variables measured, with significant correlations between several of them. All groups produced an adequate immune response, of either the innate or the adaptive type. 4. In the current study, group 2 showed the highest overall natural resistance, though none of the groups was uniformly superior with respect to all traits measured. 5. In conclusion, for reliable measurements of general immunocompetence or resistance to Salmonella, for example, it is important to measure several aspects of the immune system.     

Antibody responses to bacterial antigens in the pregnant ewe

The humoral immune response to the novel antigen Brucella abortus (S19) was examined in nonpregnant ewes and in ewes at different stages of pregnancy. Both the primary and secondary responses were monitored, by measuring the levels of total antibody, Immunoglobulin G (IgG), and complement fixing antibody. Results showed that total antibody production was significantly (p<0.01) impaired in late pregnancy and that IgG titres persisted at significantly (p<0.05) higher levels in non-pregnant animals late in the primary response and following the booster inoculation. Complement fixing antibody levels were significantly higher in non-pregnant ewes and animals inoculated post-partum. These results suggest that active immunity can be altered during pregnancy, with qualitative changes in the class and biological activity of antibody.     

Dietary β-hydroxy-β-methylbutyrate supplementation influences performance differently after immunization in broiler chickens

Dietary addition of the leucine metabolite β-hydroxy-β-methylbutyrate (HMB) promotes growth in various species. In addition, HMB is described to enhance immune responses which might be associated with metabolic costs. We elaborated further on the role of HMB in growth, metabolism and immunity of meat-type chickens using the following parameters: zootechnical performance, blood chemistry and a specific immune responses after immunization with a human serum albumin (HSA)/Freund’s (in) complete adjuvant combination. The chickens received commercial feeds either unsupplemented or supplemented with 300 mg HMB/kg feed. β-hydroxy-β-methylbutyrate-supplemented chickens were significantly heavier at 2 weeks of age but this difference was attenuated at later ages. Compared with their unsupplemented controls, cumulative feed conversion was significantly lower in HMB-supplemented chickens. There were no differences in blood chemistry between both dietary treatments. After immunization, HMB significantly attenuated the acute phase protein response at day 1 of post-immunization compared with that of their unsupplemented counterparts. After day 7 post-immunization, body weight gain of the immuno-challenged HMB-supplemented chickens was significantly depressed, but their specific anti-HSA IgG response was significantly enhanced compared with that of their immuno-challenged unsupplemented counterparts. The underlying mechanisms and signalling pathways for these phenomena need to be elucidated. Nevertheless, we are able to conclude that HMB is beneficial for performance under normal circumstances. On the other hand, HMB stimulates the immune response after an immunological challenge, though at the cost of reduced growth.     

Kinetic differences in intestinal and systemic interferon-gamma and antigen-specific antibodies in chickens experimentally infected with Eimeria maxima

Kinetic differences between systemic vs. intestinal and humoral vs. cellular immune responses were elucidated in chickens experimentally infected with Eimeria maxima by comparing interferon-gamma (IFN-gamma) and parasite-specific antibody levels in the intestine and serum during the course of infection. The level of serum IFN-gamma correlated significantly with fecal oocyst shedding (r2 = 0.97), thereby establishing the importance of cell-mediated immunity in coccidia infection. Moreover, intestinal IFN-gamma levels increased sooner than those in sera (4 vs. 6 days postinfection) and both were observed prior to the appearance of parasite-specific antibodies (8-10 days postinfection), again indicating the importance of intestinal cellular immunity in coccidiosis. Although immunoglobulin (Ig)G, IgA, and IgM isotypes of the antigen-specific antibody response increased significantly in both the intestine and serum after E. maxima infection, intestinal IgA-specific antibodies showed the most dramatic increase. However, the relevance of this observation in the context of primary Eimeria infection is unclear because the coccidia parasites have reached the final stages of their life cycle by this time. These results thus demonstrate the importance of T-cell immune responses against coccidia, characterized by local IFN-gamma secretion in the intestine, in mediating host protective immune response to coccidia.     

Comparison of adjuvants with respect to serum IgG antibody response in orally immunized chickens

We have previously shown that oral immunization with non-replicating antigens hardly induced serum IgG antibody response in chickens and addition of sodium fluoride (NaF) to the immunogen markedly improved their immunological states. In the present study, taurine, lithium and Quillaja saponin (Q-SAP) were compared with NaF with respect to their enhancement of serum IgG antibody response in chickens after oral immunization. The antibody titer of chickens which received Q-SAP as the mucosal adjuvant tended to be higher than that of chickens which received antigen plus NaF. Simultaneous administration of antigen with lithium or taurine elicited a higher antibody titer in chickens compared to those of chickens orally immunized with antigen alone, but the effect of these two adjuvants was less efficient compared with that of NaF. These results suggested that Q-SAP as well as NaF is useful as an oral adjuvant for chickens.     

The role of bursa-dependent responses in immunity to infectious laryngotracheitis.

The effect of combined day-old surgical bursectomy and cyclophosphamide therapy on the response of young chicks to Brucella abortus antigen and cloacal vaccination against infectious laryngotracheitis virus (ILTV) was examined. The combined treatment effectively prevented the development of serum antibody to both B abortus and ILTV, yet when these chickens were challenged with virulent ILTV, they were found to be immune. It is suggested that immunity to ILTV may depend on the development of a cell mediated immune response.     

Immunity to Ostertagia ostertagi

Control of ostertagiasis is a major economic problem in the temperate areas of the world. An immunological control scheme for cattle is an important aspect of integrated control and its achievement is dependent on our understanding of how immunity is acquired. Also, host mechanisms regulated by Ostertagia need to be understood in order to address immunological questions. Little is known about acquired immunity to Ostertagia infection in cattle. The degree of immunity is incomplete and its development slow. Evidence is accumulating for impairment of antibody and cellular immune responses by Ostertagia which is responsible for the survival of the parasite in the young host. Other experimental work suggests that local factors, such as eosinophil accumulation, mucosal mast cell kinetics and other cellular and humoral immune responses, may play a role in pathogenesis and immunity. At the present time these areas are also totally unexplored in the study of Ostertagia infection.     

Humoral and cell-mediated immune responses in chickens with infectious bursal disease

Primary and secondary immune responses to Newcastle disease virus (NDV) was evaluated in chickens infected with infectious bursal disease virus (IBDV) at one and 28 days of age. The geometric mean primary hemagglutination-inhibition antibody titers (GMT) of chickens infected with IBDV at one day of age was significantly lower (P less than or equal to 0.01) than those infected at 28 days of age. Infection with IBDV had no influence on secondary immune response to NDV. The effect of IBDV infection at one day of age on the cell-mediated immunity of chickens was evaluated by skin allograft acceptance or survival time. There was no significant difference between the percentage of grafts accepted in IBDV infected and noninfected control chickens. However, the mean graft survival time in the IBDV infected chickens was significantly longer (P less than or equal to 0.05) than those in the control group. This suggested a suppression of cell-mediated immunity due to IBDV infection.     

Immune response during coccidiosis in SC and FP chickens. I. In vitro assessment of T cell proliferation response to stage-specific parasite antigens

Development of cell-mediated immunity (CMI) and comparative effectiveness of different stage-specific coccidia antigens in T cell activation during avian coccidiosis were evaluated in two inbred strains of chickens using a specific in vitro T cell proliferation assay. Lymphocytes from chickens infected with different Eimeria spp. showed proliferative response to sporozoites, merozoites or Eimeria soluble antigen (Esa) excreted by cultured parasites. Detectable CMI response was observed at 21 day P.I. in chickens infected with E. tenella and E. maxima. Generally lower T cell response was observed in chickens infected with E. acervulina. Merozoites were highly immunogenic compared to sporozoites. Esa prepared from cultured parasites was as effective as whole parasites in evoking a T cell response. Although strain variation in T cell response to parasites or Esa was observed during a primary infection, substantially enhanced T cell response was observed 3 days after a secondary infection in both strains of chickens. The results of the present investigation suggest that Esa may be a major parasite antigen released to the immune system during early stages of infection and relevant to the development of protective immunity.