Genetics and vaccine efficacy: host genetic variation affecting Marek's disease vaccine efficacy in White Leghorn chickens.

Marek's disease (MD) is a T-cell lymphoma disease of domestic chickens induced by MD virus (MDV), a naturally oncogenic and highly contagious cell-associated α-herpesvirus. Earlier reports have shown that the MHC haplotype as well as non-MHC genes are responsible for genetic resistance to MD. The MHC was also shown to affect efficiency of vaccine response. Using specific-pathogen-free chickens from a series of 19 recombinant congenic strains and their 2 progenitor lines (lines 6(3) and 7(2)), vaccine challenge experiments were conducted to examine the effect of host genetic variation on vaccine efficacy. The 21 inbred lines of White Leghorns share the same B*2 MHC haplotype and the genome of each recombinant congenic strain differs by a random 1/8 sample of the susceptible donor line (7(2)) genome. Chickens from each of the lines were divided into 2 groups. One was vaccinated with turkey herpesvirus strain FC126 at the day of hatch and the other was treated as a nonvaccinated control. Chickens of both groups were inoculated with a very virulent plus strain of MDV on the fifth day posthatch. Analyses of the MD data showed that the genetic line significantly influenced MD incidence and days of survival post-MDV infection after vaccination of chickens (P<0.01). The protective indices against MD varied greatly among the lines with a range of 0 up to 84%. This is the first evidence that non-MHC host genetic variation significantly affects MD vaccine efficacy in chickens in a designed prospective study.

Relation between the major histocompatibility (B) locus and autoimmune thyroiditis in obese chickens.

Obese strain chickens develop circulating autoantibodies to thyroglobulin and lymphocytic infiltration of their thyroids during aging. Two alleles, B(1) and B(4), are found with high gene frequency at the major histocompatibility (B) locus. Greater pathology and higher antibody titers are observed in B(1)B(1) and B(1)B(4) birds than in their B(4)B(4) siblings.

Lymphoid organ size varies among inbred lines 6(3) and 7(2) and their thirteen recombinant congenic strains of chickens with the same major histocompatibility complex.

The objective was to evaluate lymphoid organ size in chickens from a series of 13 recombinant congenic strains (RCS) and their highly inbred parental lines (6(3) and 7(2)). The parental line 6(3) was selected for resistance to tumors induced by Marek's disease virus and avian leukosis viruses, whereas line 7(2) was selected for susceptibility to these tumors. Each RCS on the average contains a random one-eighth of genome from the donor line 7(2). Previous studies have shown that lines 6(3) and 7(2) differ in the size of primary lymphoid organs; i.e., the bursa of Fabricius (BF) and the lobes of the thymus (T) are smaller in line 6(3) than line 7(2). In the current study, the relative size of the T, BF, and spleen was first examined in about 15 males from each of 13 RCS and the 2 parental lines at 60 to 69 d of age. The differences of relative BF, T, and spleen size among the RCS and the parental lines 6(3) and 7(2) differed significantly (P < 0.001). Males and females from 4 RCS and the 2 parental lines were evaluated a second time, and differences in the relative sizes in lymphoid organs among the RCS and parental lines were consistent. In 2 RCS, the size of the T and BF was comparatively large as in line 7(2), leading to the conclusion that different allelic forms at 1 or more loci in these RCS regulate the size of both organs. In 2 other RCS, the BF was large compared with the T, suggesting that allelic forms at some loci in these RCS influence the BF independent of the T. The relative lymphoid organ size among the RCS appeared to cosegregate with the concentration of IgG in the plasma measured previously. The evaluation of genomic variability of these lines is underway, and the RCS are available for research on traits that differ between lines 6(3) and 7(2).

Genetic mapping of quantitative trait loci affecting susceptibility to Marek's disease virus induced tumors in F2 intercross chickens.

Marek's disease (MD) is a lymphoproliferative disease caused by the MD virus (MDV), which costs the poultry industry nearly $1 billion annually. To identify quantitative trait loci (QTL) affecting MD susceptibility, the inbred lines 6(3) (MD resistant) and 7(2) (MD susceptible) were mated to create more than 300 F2 chickens. The F2 chickens were challenged with MDV JM strain, moderately virulent) at 1 wk of age and assessed for MD susceptibility. The QTL analysis was divided into three stages. In stage 1, 65 DNA markers selected from the chicken genetic maps were typed on the 40 most MD-susceptible and the 40 most MD-resistant F2 chickens, and 21 markers residing near suggestive QTL were revealed by analysis of variance (ANOVA). In stage 2, the suggestive markers plus available flanking markers were typed on 272 F2 chickens, and three suggestive QTL were identified by ANOVA. In stage 3, using the interval mapping program Map Manager and permutation tests, two significant and two suggestive MD QTL were identified on four chromosomal subregions. Three to five loci collected explained between 11 and 23% of the phenotypic MD variation, or 32-68% of the genetic variance. This study constitutes the first report in the domestic chicken on the mapping of non-major histocompatibility complex QTL affecting MD susceptibility.

Chronic dopaminergic sensitivity after Sydenham's chorea.

We studied psychometric performance on the Minnesota Multiphasic Personality Inventory (MMPI) mini-mult and drug-induced choreic reactions in a group of patients with a history of Syndenham's chorea. Action tremor, motor signs, and residual chorea were common. One-half of the patients reported adverse choreic reactions to one or more agents. Patients with adverse reactions to central stimulants and anorectics had statistically significant elevations in the psychotic tetrad of the MMPI. Sydenham's chorea in childhood seems to confer persistent sensitivity to agents that augment central dopaminergic activity, which may be expressed as acute chorea. Central dopaminergic sensitivity may explain earlier reports of psychologic difficulties in survivors of rheumatic chorea.

PIP: 32 patients with Sydenham's chorea were studied at the La Rabida Institute for psychometric performance on the Minnesota Multiphasic Personality Inventory (MMPI). Questionnaires used included a definition of chorea and a description of choreic movements which the patients and members of their households were asked to read. Results were: 1) the only medical condition frequently reported was arthritis; 20/32 patients reported medical consultation for this complaint; 2) 19 patients including 2 with chorea gravidium, reported motor or psychiatric side effects from 1 or more agents; 3) in patients with multiple drug exposures a history of adverse motor reactions to decongestants was always associated with adverse reactions to anorectics or amphetamine in patients with exposure to all agents, and a similar pattern was noted with thyroid hormone and oral contraceptives (OCs); 4) 1 patient with chorea gravidium reported dyskinesias after administration of decongestants or amphetamine but tolerated OCs; and 5) MMPI scores from patients reporting adverse responses to amphetamines were statistically elevated in the psychotic tetrad. This study provides support for the belief that Sydenham's chorea is not a benign self-limited disease of childhood. In addition to mild residual neurologic abnormalities, the disorder appears to confer long-standing sensitivity to a variety of dopaminergically active agents.

Facial recognition memory in dementia.

Previous investigations of memory in senile dementia of the Alzheimer's type (SDAT) have focused on verbal learning and memory. The aim of the present study was to determine whether the amnesia of SDAT is limited to verbal material. Patients with SDAT (N = 29; mean age = 69.3) and healthy normal controls (N = 41; mean age = 69.3) were given a test of facial perception and two recognition memory tasks, one for words and one for faces. The results indicate that dementia patients show a deficit in the retention of facial information. This deficit cannot be attributed to faculty initial perception or to a response bias. The verbal and facial memory deficits in SDAT appear to differ: performance on tests of verbal and facial memory is relatively independent, and substantial encoding and linguistic defects contribute to the verbal, but not the facial, memory disorder resulting in more severe impairment on tests of verbal memory. The implications of these findings for research on the neuropharmacology and pathophysiology of SDAT are discussed.

Bursa-independent origin of Fc IgG receptor-bearing lymphocytes in chickens.

On the basis of performed studies the following observations indicate that there is a bursa-independent origin of lymphocytes with Fc IgG receptors: (a) the first appearance of Fc positive cells is in fetal liver and bone marrow, (b) a relative low percentage of Fc receptor cells, as compared to B cells, occurs in the bursa during the embryonic and post-hatching period, (c) a far lower percentage of Fc positive cells than B cells is found in the spleen, (d) a normal level of Fc positive cells occurs in bursectomized birds, (e) a majority of these cells do not possess thymic or bursa antigen or surface immunoglobulin.

Detection of endogenous avian leukosis virus envelope in chicken plasma using R2 antiserum.

Immunologic tolerance to oncogenic avian leukosis virus (ALV) is mediated, in part, by the interaction of endogenous ALV (EV) envelope and immune competent cells. A flow cytometry method is described for detecting the EV envelope in chicken plasma or serum. The method employs two types of target red blood cells (RBC) obtained from chickens lacking EV; RBC susceptible to EV infection (containing EV receptors), and those resistant to EV infection (lacking EV receptors). RBC from susceptible chickens will bind EV envelope glycoprotein (gp85) when present in plasma. The gp85-bound RBC are subsequently incubated with a highly specific chicken alloantibody, termed R2. Using flow cytometry, gp85 is detected indirectly with a fluoresceine-tagged antibody to chicken immunoglobulin; plasmas lacking gp85 are nonreactive and fluorescence remains at a background level. Because RBC from resistant chickens are nonreactive regardless of the presence or absence of EV gp85, a specific binding index was calculated to compare relative binding of EV gp85 on susceptible and resistant RBC, and thus identify chickens that express EV gp85. The specificity of the assay was demonstrated using plasma from chickens of 14 standard laboratory lines previously defined for EV envelope expression including two sets of highly congenic lines that differ in EV expression. This assay detects differences attributable to EV gp85 in chickens of commercial breeding lines of White Leghorns and broilers. Moreover, if chickens lack EV, the R2 plasma assay can differentiate between EV-susceptible and EVresistant siblings.

Characterization and experimental reproduction of peripheral neuropathy in White Leghorn chickens.

A clinical neurological syndrome termed peripheral neuropathy (PN) that resembles Marek's disease (MD) occurred at low frequency in a commercial layer strain for several years. Study of chickens from six field cases showed that the PN syndrome could be distinguished pathologically from MD on the basis of several factors, including onset as early as 6 weeks, presence of B-type but not A-type lesions in peripheral nerves, and absence of visceral lymphomas. Serotype 1 MD virus could not be isolated from blood from any chicken or demonstrated in tissues by histochemistry or polymerase chain reaction assays. Moreover, the syndrome was not prevented by MD vaccination, either in the field or in laboratory trials. PN was induced in 3 to 54%of commercial line chickens inoculated at 1 or 6 days of age with whole blood or buffy coat cells from clinically affected donor chickens. Sonicated cells also induced PN, but plasma was ineffective. Chickens did not develop PN if reared in isolators without cellular transfer or when vaccinated solely against MD. However, PN was observed in 9% of 57 B*2/*19 commercial chickens reared in isolators following vaccination against MD, infectious bursal disease, Newcastle disease and infectious bronchitis, suggesting that common vaccines may predispose chickens to PN. The data confirmed a strong influence of the major histocompatibility complex (B-complex) on both naturally occurring and experimentally induced PN with the B*19 haplotype conferring susceptibility compared with other alleles. It is postulated that PN may represent an autoimmune reaction to nerve tissue that may result from response to a combination of common vaccines. These studies confirmed that PN is distinct from MD, provided criteria for its differential diagnosis, identified strategies for its control, and established a model for its experimental induction.

B-congenic chickens differ in macrophage inflammatory responses.

The influence of the chicken major histocompatibility (B) complex (MHC) on monocyte and macrophage recruitment and activation was examined using fully developed 15I5-B congenic White Leghorn lines (ten backcross generations). The phagocytic activity of Sephadex-elicited peritoneal macrophages for sheep red blood cells (SRBCs) was highest in lines 15.7-B2 and 15.P-B13 and lowest in 15.15I-B5 and 15.N-B21. The same pattern of phagocytic activity was obtained when LPS (E. coli) was used as the in vivo elicitor-activator of peritoneal macrophages. Lines with B2 and B13 haplotypes had elevated percentages of phagocytic macrophages and a higher internalization activity per cell than did B5 and B21 congenic chickens. Differential peritoneal macrophage function between congenic lines was further supported by quantitation of superoxide anion release. B2 and B13 haplotypes were associated with high activity in contrast with B5, which was low, and 15I5 (B15) and B21 which were intermediate for superoxide anion release by macrophages. In vitro activation of blood monocytes with LPS resulted in similar line differences for SRBC phagocytic activity as were observed with in vivo Sephadex and LPS activation. In contrast, chemotaxis of blood mononuclear leukocytes to f-met-leu-phe produced a reciprocal response pattern among the haplotypes. Cells from lines with haplotypes B5 and B21 were superior to those of B2, B13, and B15 congenic lines in their directed migration towards this chemoattractant. All functional differences occurred despite similarities among lines in the cellular profiles of both elicited peritoneal exudate cells and isolated blood mononuclear cells.

Chemotactic activity of chicken blood mononuclear leukocytes from 15I5-B-congenic lines to bacterially-derived chemoattractants.

The chemotactic activity of chicken blood mononuclear leukocytes was examined in partially-developed 15I5-B-congenic chicken lines using Enterobacter cloacae culture supernatant and formyl-methionyl-leucyl-phenylanine (f-met-leu-phe) (10(-5)M). Cells from seven different coded B-congenic lines were used to study each chemoattractant in vitro. Mononuclear cells from lines .15I-B5,.C-B12 and the background line 15I5 (B15) exhibited a significantly greater directed migration to bacterial supernatant than did cells from four lines carrying the B2, B2, B13, and B19 haplotypes, respectively. Similarly response to f-met-leu-phe was greatest in lines .15I-B5,.C-B12 and .N-B21, with the same four lines exhibiting a significantly lower response. Since f-met-leu-phe was originally isolated from bacteria, the results indicate that these lines possess differential chemotactic responses to certain bacterially-derived chemoattractants. Major histocompatibility complex (MHC) differences between the lines may serve as a genetic basis for the differential responses. Extrapolation of these results to other chemotactic-receptor systems would require further examination.

Genetic control of immunity to Eimeria tenella. Interaction of MHC genes and non-MHC linked genes influences levels of disease susceptibility in chickens.

The relative importance of MHC genes and background genes in the genetic control of disease susceptibility and the development of protective immunity to E. tenella infection was investigated in eight different strains of 15I5-B congenic and four inbred chicken strains. RPRL 15I5-B congenic chickens that share a common genetic background but express different B haplotypes demonstrated wide variations in disease susceptibility and the development of acquired resistance to E. tenella infection. Infection of chickens sharing a common B haplotype but expressing different genetic backgrounds showed quite contrasting levels of susceptibility to secondary E. tenella infection. In all chicken strains examined, infected chickens developed high levels of serum and biliary anti-coccidial antibodies regardless of their B haplotypes. Furthermore, no correlation between antibody levels and the phenotypically expressed levels of disease resistance was demonstrated. These findings lend support to the view that interaction of MHC genes and non-MHC genes influences the outcome of host response to E. tenella infection.

Influence of B-haplotype on the relative efficacy of Marek's disease vaccines of different serotypes.

To determine if B-haplotype differentially influences vaccinal immunity to very virulent Marek's disease (MD) virus challenge, chickens of five 15.B-congenic lines were vaccinated with vaccines representing serotypes 1, 2, and 3. B-haplotype differentially influenced vaccinal immunity to very virulent MD virus challenge using vaccines of all three serotypes, and different MD vaccines were optimal for some B-haplotypes. Regarding specific haplotypes, the 15.B-congenic chickens with B2 and B13 developed less protection against MD than chickens with B15 following vaccination with all three serotypes of MD vaccine, whereas the chickens with B5 and B21 developed variable protection with different MD vaccines. Thus, for induction of maximum MD resistance, it may be necessary to select a vaccine appropriate for the predominant B-haplotypes of the chicken flock.

Serotype specificity of B-haplotype influence on the relative efficacy of Marek's disease vaccines.

B-haplotype genes in the chicken were previously shown to differentially influence vaccine efficacy against challenge with very virulent Marek's disease virus according to the type of Marek's disease (MD) vaccine used. To determine whether MD vaccines of the same serotype gave comparable levels of protection against MD in chickens of the same haplotype challenged with MD virus strain Md5, two serotype 1 and two serotype 2 vaccines were compared with one serotype 3 vaccine using chickens of 15-B-congenic lines. There was a strong correlation in development of MD lesions among chickens of the different lines receiving the two serotype 2 vaccines (r = 0.94) as well as among chickens receiving the two serotype 1 vaccines (r = 0.76). The serotype 1 vaccines were preferable for B2, B13, B15, and B21, but serotype 2 vaccines were more protective for B5 chickens. The two serotype 2 vaccines gave equivalent protection; however, of the serotype 1 vaccines, CVI988/Rispens provided more protection than Md11/75c/R2/23. We conclude that the B-haplotype influence on MD vaccine efficacy is dependent on the serotype of the vaccine.

Influence of turkey herpesvirus vaccination on the B-haplotype effect on Marek's disease resistance in 15.B-congenic chickens.

Eight recently developed 15.B congenic lines of chickens were tested for Marek's disease (MD) resistance by intra-abdominal injection of cell-associated preparations of MD virus of a virulent strain (JM), a very virulent strain (Md5), or Md5 after vaccination with turkey herpesvirus (HVT) strain FC126. Chickens of the 15.N congenic line (B15B21 or B21B21) were very resistant to JM-induced MD, in contrast to chickens homozygous for the B-haplotypes 2, 5, 12, 13, 15, or 19. After Md5 infection, more than 88% of the chickens in all of the congenic lines developed MD. However, when chickens were vaccinated with HVT before being inoculated with Md5, the B5 and B12 homozygotes were more resistant to MD than were the B2, B13, or B19 homozygotes, and B15 and B21 homozygotes had intermediate resistance. B5B5 and B2B5 F2 chicks inoculated with HVT and Md5 had a lower prevalence of MD than B2B2 sibs. These results demonstrate that a protocol involving HVT vaccination of chicks followed by infection with very virulent MD virus will allow the detection of B-haplotypes determining MD resistance, some of which are not detectable in unvaccinated chicks challenged with virulent MD.

Response of B congenic chickens to infection with infectious bursal disease virus.

Six congenic lines of chickens that differ from the parental inbred line RPRL-15I5 for genes in the major histocompatibility (B) complex were used to study the influence of the B haplotypes on the response of chickens to infection with virulent infectious bursal disease virus (IBDV) at 1 day or 4 weeks of age, and on the antibody response to vaccination with live or inactivated oil-emulsion (OE) IBDV vaccines at 7 weeks of age. IBDV-induced immunodepression and lesions in the bursa, spleen, and thymus in chickens infected with virus at 1 day of age were of the same degree of severity, regardless of line of chickens used. The response of blood cells to the mitogens phytohemagglutinin-M and concanavalin A was elevated in chickens infected with IBDV at 1 day of age. In an experiment conducted to study the effect of the B haplotype on IBDV infection in 4-week-old chickens, B congenic line C-12 (B12B12) showed the highest susceptibility to clinical IBD, with mortality of 79%. No detectable difference in the serological response to vaccination with live or OE IBDV vaccines was noted among chickens of various congenic lines. We conclude that the B haplotypes may influence IBDV-induced mortality, but not immunodepression or severity of lesions in lymphoid organs, or the antibody response to live or OE IBDV vaccines.

Increase of intestinal intraepithelial lymphocytes expressing CD8 antigen following challenge infection with Eimeria acervulina.

Eimerian infection-induced changes in the intestinal intraepithelial lymphocyte (IEL) subpopulations expressing CD8 antigen (cytotoxic/suppressor T cells) or antigen-specific T cell receptor (TCR) heterodimer alpha beta (TCR2) or gamma delta (TCR1) were investigated in F2 crosses of 15I5 B-congenic chickens differing for the major histocompatibility complex (MHC). Duodenum TCR2+ IEL were increased in B2 B2 and B5 B5 chickens 7 days following secondary infection. Two-color immunofluorescence revealed that the majority of CD8+ cells in the duodenum intraepithelium of immune chickens expressed TCR2. A significant increase in the duodenum TCR2+CD8+ and TCR1+CD8+ IEL occurred in B2 B2 chickens, which developed significantly less oocyst production than the B5 B5 chickens following challenge infection. The results suggest that a significant increase in the duodenum CD8+ IEL may reflect an enhanced acquired immunity of B2 B2 chickens.

Mitogen-induced lymphocyte proliferation and interferon production following coccidia infection.

Concanavalin A-induced lymphocyte proliferation and interferon production were measured following infection with Eimeria acervulina in lines of chickens congenic at the major histocompatibility complex and in two unrelated lines. Similar proliferation responses were seen for splenic and peripheral blood lymphocytes following primary but not secondary infection. Greater differences in lymphocyte proliferation were found between the congenic lines and unrelated lines than among the congenic lines. The congenic lines had very low responses to Con A, in terms of both lymphocyte proliferation and interferon production. At some times after infection, background proliferation in all lines was higher for infected than uninfected chickens, which was probably an indication of activated lymphocytes in the circulation. Mitogen-induced lymphocyte proliferation and interferon production, although clearly affected by coccidial infection, were not reflective of severity of infection.

Partial inhibition by turkey herpesvirus of serotype 2 Marek's disease virus plaque formation and in vivo infectivity.

The increased use of serotype 2 Marek's disease virus (MDV) and serotype 3 turkey herpesvirus (HVT) as components of effective bivalent vaccines against Marek's disease (MD) prompted studies on the possible interactions of these two viruses in vitro and in vivo. The replication of the SB-1 strain of MDV was compared with replication of the FC126/2 strain of HVT in chickens and cell cultures infected with one or both viruses. Replication of MDV was reduced in the presence of HVT in both in vitro and in vivo systems. MDV plaque counts in dually infected chicken embryo fibroblast cultures inoculated with tissue-culture-propagated viruses were reduced by up to 91%; however, no inhibition was noted when inocula consisted of virus-infected buffy-coat cells. Plaque formation by MDV in chicken embryo fibroblast cultures was inhibited by virus-free conditioned medium from HVT-infected cultures. This conditioned medium also inhibited growth of vesicular stomatitis virus in a standard interferon assay. In chickens inoculated with both MDV and HVT, MDV viremia titers were lower and the dose required to infect 50% of susceptible chickens was increased 13-fold compared with chickens inoculated with MDV alone. In spite of these findings, there was no evidence that high concentrations of HVT interfered with either the ability of MDV to induce protective synergism in vivo or the protective efficacy of bivalent vaccines. No reciprocal inhibitory effects of MDV on the replication of HVT in vivo or in vitro were noted.

Absence of influence on immune competence by the sex-linked gene (K) determining slow feathering in white leghorn chickens.

Males of white leghorn strain crosses heterozygous (Kk) for the sex-linked feathering locus genes were mated to rapid-feathering (k-) females to produce rapid- and slow-feathering chicks of both sexes. K did not influence humoral-mediated immunity against challenge with sheep erythrocytes, killed Brucella abortus, or killed infectious bursal disease virus. Chicks challenged at 3 weeks of age had higher primary responses and higher titers of 2-mercaptoethanol-resistant antibody (IgG) than those challenged at 1 week of age. K had no influence on levels of cell-mediated immunity based on responses in in vitro phytohemagglutinin tests of donors 2 to 126 days of age, on responses in in vitro one-way mixed-lymphocyte cultures, and on rejection rates of skin grafts on young chicks. Feathering type did not influence viremia or antibody to avian leukosis virus; the level of lymphoid leukosis tumors was higher in rapid-feathering females than in slow-feathering females at 28 weeks of age (53% vs. 72%; P less than or equal to 0.10). We conclude that K does not influence general immune competence. The possibility that it may influence specific immunity to ALV under conditions not met in this study, because of an endogenous virus recently associated with the K locus, is discussed.