Antibody Repertoire Development in Swine.

We describe the domestication of the species, explore its value to agriculture and bioscience, and compare its immunoglobulin (Ig) genes to those of other vertebrates. For encyclopedic information, we cite earlier reviews and chapters. We provide current gene maps for the heavy and light chain loci and describe their polygeny and polymorphy. B-cell and antibody repertoire development is a major focus, and we present findings that challenge several mouse-centric paradigms. We focus special attention on the role of ileal Peyer's patches, the largest secondary lymphoid tissues in newborn piglets and a feature of all artiodactyls. We believe swine fetal development and early class switch evolved to provide natural secretory IgA antibodies able to prevent translocation of bacteria from the gut while the bacterial PAMPs drive development of adaptive immunity. We discuss the value of using the isolator piglet model to address these issues.

Antibody repertoire development in fetal and neonatal piglets. XXIV. Hypothesis: The ileal Peyer patches (IPP) are the major source of primary, undiversified IgA antibodies in newborn piglets.

The ileal Peyers patches (IPP) of newborn germfree (GF) piglets were isolated into blind loops and the piglets colonized with a defined probiotic microflora. After 5 weeks, IgA levels in the intestinal lavage (IL) of loop piglets remained at GF levels and IgM comprised ∼70% while in controls, IgA levels were elevated 5-fold and comprised ∼70% of total Igs. Loop piglets also had reduced serum IgA levels suggesting the source of serum IgA had been interrupted. The isotype profile for loop contents was intermediate between that in the IL of GF and probiotic controls. Surprisingly, colonization alone did not result in repertoire diversification in the IPP. Rather, colonization promoted pronounced proliferation of fully switched IgA(+)IgM(-) B cells in the IPP that supply early, non-diversified "natural" SIgA antibodies to the gut lumen and a primary IgA response in serum.

Antibody repertoire development in fetal and neonatal piglets. XII. Three IGLV genes comprise 70% of the pre-immune repertoire and there is little junctional diversity.

We characterized 239 lambda rearrangements from fetal and germfree (GF) piglets to: (1) determine if transcripts recovered from the earliest sites of B cell lymphogenesis were unique (2) determine what proportion of the genome is used to form the pre-immune repertoire (3) estimate the degree of somatic hypermutation and junctional diversity during ontogeny and (4) test whether piglets maintained germfree in isolators (GF piglets) have a more diversified repertoire than fetal piglets. We show that all expressed lambda genes belong to the IGLV3 and IGLV8 families and only IGLJ2 and IGLJ3 were expressed and used equally throughout fetal and neonatal life. Only genes of the IGLV8 family were used in yolk sac and fetal liver and in these tissues, IGLV8-10 comprised >50%. However, the IGLV8 genes recovered at these early sites of B cell lymphogenesis were recovered at all stages of development. Thus, no unique lambda rearrangement was recovered at the first sites of B cell development. The frequency of somatic hypermutation (SHM) in fetal piglets was ~5.9 per Kb equivalent, mutation were concentrated in CDR regions and did not increase in GF piglets. The average CDR3 length was 30 nt ± 2.7 and did not change in GF piglets. Similar to the heavy chain pre-immune repertoire in this species, three IGLV genes account for ~70% of the repertoire. Unlike the heavy chain repertoire, junctional diversity was very limited.

Antibody repertoire development in fetal and neonatal piglets. XIV. Highly restricted IGKV gene usage parallels the pattern seen with IGLV and IGHV.

Kappa transcripts from fetal piglets were compared to the recently reported kappa genome. Although five IGKV gene families are present in the genome, only IGKV1 and IGKV2 family genes are transcribed; the latter comprises >95% of the repertoire, in which two genes account for ~80%. We provisionally identified a new sequence allele of IGKV2-10 and two new IGKV genes that were not present in the genome of a single Duroc sow. One of these (IGKV2-1) accounted for 39% of the total pre-immune repertoire. The discovery of new IGKV genes and alleles in only 90 transcripts from mixed breeds, suggests considerable polymorphism and polygeny in the kappa locus of swine. Similar to lambda rearrangements, CDR3 length and diversity is restricted. The somatic mutation frequency is low and accumulates in especially CDR1. This transcriptional analysis of the pre-immune kappa repertoire completes a comparative study of all three Ig loci which has allowed the potential and actual combinatorial repertoire to be determined. Calculations show that combinatorial diversity in all three loci contribute comparatively little to the swine pre-immune antibody repertoire. Compared to humans that can potentially generate a million binding site variants, only 16-48 variant comprise 70% of the swine repertoire and 224 account for 95-100%. The frequency of somatic mutation does not differ among rearrangements from all three loci and the CDR3 diversity index shows that swine overwhelmingly generate their pre-immune repertoire by junctional diversity in heavy chain rearrangements.

Antibody repertoire development in fetal and neonatal piglets. XVI. Influenza stimulates adaptive immunity, class switch and diversification of the IgG repertoire encoded by downstream Cγ genes.

Infection of germ-free isolator piglets with swine influenza (S-FLU) that generates dsRNA during replication causes elevation of immunoglobulins in serum and bronchoalveolar lavage, a very weak response to trinitrophenyl conjugates but an immune response to S-FLU. The increased immunoglobulin levels result mainly from the polyclonal activation of B cells during the infection, but model antigen exposure may contribute. The 10-fold increase in local and serum IgG accompanies a 10-fold decrease in the transcription of IgG3 in the tracheal-bronchial lymph nodes and in the ileal Peyer's patches. Infection results in class switch recombination to downstream Cγ genes, which diversify their repertoire; both features are diagnostic of adaptive immunity. Meanwhile the repertoires of IgM and IgG3 remain undiversified suggesting that they encode innate, natural antibodies. Whereas IgG3 may play an initial protective role, antibodies encoded by downstream Cγ genes with diversified repertoires are predicted to be most important in long-term protection against S-FLU.

Antibody repertoire development in fetal and neonatal piglets. XV. Porcine circovirus type 2 infection differentially affects serum IgG levels and antibodies to ORF2 in piglets free from other environmental factors.

Porcine circovirus type 2 (PCV2) is an important pathogen in the porcine respiratory disease complex (PRDC) and its persistence may be due to dysregulation of systemic immunity. We examined this contention using isolator piglets. We present data on Ig levels in serum and bronchio-alveolar lavage (BAL), on antibody response to PCV2 and to TNP conjugates used as model antigens in 48 PCV2-infected isolator piglets. We compared these to data from TNP-immunized isolator piglets colonized with a probiotic flora, those infected with swine influenza (S-FLU) and those infected with porcine respiratory and reproductive syndrome virus (PRRSV). We found that PCV2 infection does not cause generalized hypergammaglobulinemia that characterizes PRRSV infections, but causes an unexplained increase in serum IgA. All animals had serum IgG to the ORF2 gene product of PCR2, but neither IgA nor IgG anti-ORF2 responses in BAL. PCV2 infection is a poor adjuvant since only natural anti-TNP antibodies were found. Unexpectedly, immunization appeared to result in lower Ig levels and lower anti-ORF2 responses. There was extreme variation in serum Ig levels in response to infection that could in part be traced to genetic and gender differences. These data suggest that non-replicating vaccines are unlikely to result in a significant primary antibody response but may prime the system for a secondary antibody and cytotoxic response following actual infection. In any case, developers may have to contend with significant genetic differences in the response of piglets to PCV2.

The porcine antibody repertoire: variations on the textbook theme.

The genes encoding the heavy and light chains of swine antibodies are organized in the same manner as in other eutherian mammals. There are ∼30 VH genes, two functional DH genes and one functional JH gene, 14-60 Vκ genes, 5 Jκ segments, 12-13 functional Vλ genes, and two functional Jλ genes. The heavy chain constant regions encode the same repertoire of isotypes common to other eutherian mammals. The piglet models offers advantage over rodent models since the fetal repertoire develops without maternal influences and the precocial nature of their multiple offspring allows the experimenter to control the influences of environmental and maternal factors on repertoire development postnatally. B cell lymphogenesis in swine begins in the fetal yolk sac at 20 days of gestation (DG), moves to the fetal liver at 30 DG and eventually to the bone marrow which dominates until birth (114 DG) and to at least 5 weeks postpartum. There is no evidence that the ileal Peyers patches are a site of B cell lymphogenesis or are required for B cell maintenance. Unlike rodents and humans, light chain rearrangement begins first in the lambda locus; kappa rearrangements are not seen until late gestation. Dissimilar to lab rodents and more in the direction of the rabbit, swine utilize a small number of VH genes to form >90% of their pre-immune repertoire. Diversification in response to environmental antigen does not alter this pattern and is achieved by somatic hypermutation (SHM) of the same small number of VH genes. The situation for light chains is less well studied, but certain Vκ and Jκ and Vλ and Jλ are dominant in transcripts and in contrast to rearranged heavy chains, there is little junctional diversity, less SHM, and mutations are not concentrated in CDR regions. The transcribed and secreted pre-immune antibodies of the fetus include mainly IgM, IgA, and IgG3; this last isotype may provide a type of first responder mucosal immunity. Development of functional adaptive immunity is dependent on bacterial MAMPs or MAMPs provided by viral infections, indicating the importance of innate immunity for development of adaptive immunity. The structural analysis of Ig genes of this species indicate that especially the VH and Cγ gene are the result of tandem gene duplication in the context of genomic gene conversion. Since only a few of these duplicated VH genes substantially contribute to the antibody repertoire, polygeny may be a vestige from a time before somatic processes became prominently evolved to generate the antibody repertoire. In swine we believe such duplications within the genome have very limited functional significance and their occurrence is therefore overrated.

Antibody repertoire development in fetal and neonatal piglets. XXII. λ Rearrangement precedes κ rearrangement during B-cell lymphogenesis in swine.

VDJ and VJ rearrangements, expression of RAG-1, Tdt and VpreB, and the presence of signal joint circles (SJC) were used to identify sites of B-cell lymphogenesis. VDJ, VλJλ but not VκJκ rearrangements or SJC were recovered from yolk sac (YS) at 20 days of gestation (DG) along with strong expression of VpreB and RAG-1 but weak Tdt expression. VλJλ rearrangements but not VκJκ rearrangements were recovered from fetal liver at 30-50 DG. SJC were pronounced in bone marrow at 95 DG where VκJκ rearrangements were first recovered. The VλJλ rearrangements recovered at 20-50 DG used some of the same Vλ and Jλ segments seen in older fetuses and adult animals. Hence the textbook paradigm for the order of light-chain rearrangement does not apply to swine. Consistent with weak Tdt expression in early sites of lymphogenesis, N-region additions in VDJ rearrangements were more frequent at 95 DG. Junctional diversity in VλJλ rearrangement was limited at all stages of development. There was little evidence for B-cell lymphogenesis in the ileal Peyer's patches. The widespread recovery of VpreB transcripts in whole, non-lymphoid tissue was unexpected as was its recovery from bone marrow and peripheral blood monocytes. Based on recovery of SJC, B-cell lymphogenesis continues for at least 5 weeks postpartum.

Structure of the genomic sequence comprising the immunoglobulin heavy constant (IGHC) genes from Sus scrofa.

The number of immunoglobulin heavy chain (IGH) constant genes (IGHC) varies among mammals. To annotate the porcine IGHC genes, we sequenced the entire IGHC-containing genomic region from a single porcine haplotype. The resulting contiguous sequence included in 5' the IGH diversity (D) gene cluster and in 3' TMEM121, which flank the IGHC cluster in the human genome, suggesting that we had obtained the entire genomic region containing porcine IGHC. This region was about 190-kb long, in good agreement with those of other mammals. The porcine IGHC cluster contained 10 genes, IGHM, IGHD, six IGHG genes, IGHE and IGHA. The porcine IGHG genes formed a cluster between IGHD and IGHE, with IGHG3 considered as the most ancient IGHG gene, located at the beginning of the IGHG cluster. Furthermore, the porcine sequence contained two IGHG5 and two IGHG6 genes, but no IGHG genes for IgG2 and IgG4, suggesting flexibility within the IGHG cluster. We also recorded structural differences in the switch regions of the IGHC genes that may be important in their transcription. This haplotype can serve as a reference for future studies on other haplotypes and for functional analysis of porcine immunoglobulin (IG) isotypes.

Antibody repertoire development in fetal and neonatal piglets XXI. Usage of most VH genes remains constant during fetal and postnatal development.

Usage of variable region gene segments during development of the antibody repertoire in mammals is unresolved in part because of the complexity of the locus in mice and humans and the difficulty of distinguishing intrinsic from extrinsic influences in these species. We present the first vertical studies on VH usage that spans the fetal and neonatal period using the piglet model. We tracked VH usage in DNA rearrangements and in VDJ transcripts throughout 75 days of gestation (DG) in outbred fetuses, thereafter in outbred germfree and colonized isolator piglets, isolator piglets infected with swine influenza and in conventionally reared nematode-infected adults. Seven VH genes account for >90% of the pre-immune repertoire which is the same among tissues and in both transcripts and DNA rearrangements. Statistical modeling supports the view that proportional usage of the major genes remains constant during fetal life and that postnatal usage ranking is similar to that during fetal life. Changes in usage ranking are developmental not antigen dependent. In this species exposure to environmental antigens results in diversification of the repertoire by somatic hypermutation of the same small number of VH genes that comprise the pre-immune repertoire, not by using other VH gene available in the germline. Therefore in swine a small number of VH genes shape the antibody repertoire throughout life questioning the need for extensive VH polygeny.

Antibody repertoire development in fetal and neonatal piglets. XX. B cell lymphogenesis is absent in the ileal Peyer's patches, their repertoire development is antigen dependent, and they are not required for B cell maintenance.

The continuous ileal Peyer's patches (IPP) of sheep are regarded as a type of mammalian bursal equivalent where B cells diversify their repertoire in an Ag-independent fashion. Anatomically and developmentally similar IPP occur in swine. Resection of ∼90% of the IPP in piglets at birth did not alter Ig levels in serum and secretions or retard diversification of the Ab repertoire when animals were maintained in isolators and colonized with a defined gut flora. Resection or sham surgery elevated IgG and IgA in serum and in lavage fluid from the gut, lung, and in saliva. No changes in the frequency of IgG-, IgA-, and IgM-containing cells in the spleen and peripheral lymph node were observed. Using an index that quantifies diversification of the VDJ repertoire, no differences were seen in three secondary lymphoid tissues between piglets lacking IPP and colonized controls, whereas both groups displayed >10-fold greater diversification than did late-term fetal piglets or piglets maintained germ-free. Somatic hypermutation was very low in fetal IPP and the IPP of germ-free piglets but increased 3- to 5-fold after colonization. D-J signal joint circles were not recovered in IPP, and V-DJ signal joint circles were 5-fold lower than in bone marrow and similar to those in thymus and spleen. We conclude that the porcine IPP are not a site of B cell lymphogenesis, do not undergo Ag-independent repertoire diversification, and are not primary lymphoid tissue since they are not required for maintenance of Ig levels in serum and secretions.

The two suborders of chiropterans have the canonical heavy-chain immunoglobulin (Ig) gene repertoire of eutherian mammals.

Bats comprise 20% of all mammals, yet little is known about their immune system and virtually nothing about their immunoglobulin genes. We show that four different bat species transcribe genes encoding IgM, IgE, IgA and IgG subclasses, the latter which have diversified after speciation; the canonical pattern for eutherian mammals. IgD transcripts were only recovered from insectivorous bats and were comprised of CH1, CH3 and two hinge exons; the second hinge exon was fused to CH3. IgA in all species resembles human IgA2 with the putative cysteine forming the bridge to the light chain found at position 77. Sequence comparisons yielded no evidence for a diphyletic origin of the suborders. Bats show no close similarity to another mammalian order; the strongest association was with carnivores. Data reveal that CH diversity and VDJ and CDR3 organization are similar to other eutherian mammals, although the expressed VH3 family repertoire was unusually diverse.

The little brown bat, M. lucifugus, displays a highly diverse V H, D H and J H repertoire but little evidence of somatic hypermutation.

Myotis lucifugus populations in Northeastern US are being decimated by a fungal disease. Since almost nothing is known about the immune system of bats, we are characterizing the immunoglobulin genes of bats. We show that M. lucifugus has a diverse V(H) gene repertoire comprised of five of the seven human V(H) gene families and an estimated 236V(H)3 genes. 95% of these germline VH3 genes differ in FR3. A comparison of 67 expressed V(H)3 genes with 75 germline V(H)3 genes revealed a mutation frequency similar to fetal piglets never exposed to environmental antigens. Analysis of CDR3 regions identified at least 13 putative J(H) segments and a large D(H) repertoire. The low mutation frequency, highly diverse V(H), D(H), and J(H) germline repertoire suggests that this species may rely more on combinatorial and junctional diversity than on somatic hypermutation raising questions about the ability of M. lucifugus to respond rapidly to emerging pathogens.

Antibody repertoire development in fetal and neonatal piglets. XI. The relationship of variable heavy chain gene usage and the genomic organization of the variable heavy chain locus.

In this study, we have mapped the 3' H chain V region (V(H)) genes and those in the H chain diversity, H chain joining, and 5' portion of the H chain constant locus. We show that swine possess only two functional H chain diversity segments and only one functional H chain joining segment. These data help to explain more than a decade of observations on the preimmune repertoire of this species and reveal the vulnerability of swine to natural or designed mutational events. The results are consistent with earlier studies on the region containing Enh, Cmu, and Cdelta while revealing that the ancestral IgG3 is the most 5' Cgamma gene. We also observed a recent duplication ( approximately 1.6 million years ago) in the V(H) locus that contains six of the seven V(H) genes that comprise 75% of the preimmune repertoire. Because there are no known transfers of immune regulators or Ags that cross the placenta as in mice and humans, fetal V(H) usage must be intrinsically regulated. Therefore, we quantified V(H) usage in fetal piglets and demonstrated that usage is independent of the position of V(H) genes in the genome; the most 3' functional V(H) gene (IGHV2) is rarely used, whereas certain upstream genes (IGHV14 and IGHV15) are predominately used early in fetal liver but seldom thereafter. Similar to previous studies, three V(H) genes account for 40% of the repertoire and six for approximately 70%. This limited combinatorial diversity of the porcine V(H) repertoire further emphasizes the dependence on CDR3 diversity for generating the preimmune Ab repertoire of this species.

Porcine IgG: structure, genetics, and evolution.

Eleven genomic porcine Cgamma gene sequences are described that represent six putative subclasses that appear to have originated by gene duplication and exon shuffle. The genes previously described as encoding porcine IgG1 and IgG3 were shown to be the IgG1(a) and IgG1(b) allelic variants of the IGHG1 gene, IgG2a and IgG2b are allelic variants of the IGHG2 gene, while "new" IgG3 is monomorphic, has an extended hinge, is structurally unique, and appears to encode the most evolutionarily conserved porcine IgG. IgG5(b) differs most from its putative allele, and its C(H)1 domain shares sequence homology with the C(H)1 of IgG3. Four animals were identified that lacked either IgG4 or IgG6. Alternative splice variants were also recovered, some lacking the C(H)1 domain and potentially encoding heavy chain only antibodies. Potentially, swine can transcribe >20 different Cgamma chains. A comparison of mammalian Cgamma gene sequences revealed that IgG diversified into subclasses after speciation. Thus, the effector functions for the IgG subclasses of each species should not be extrapolated from "same name subclasses" in other species. Sequence analysis identified motifs likely to interact with Fcgamma receptors, FcRn, protein A, protein G, and C1q. These revealed IgG3 to be most likely to activate complement and bind FcgammaRs. All except IgG5(a) and IgG6(a) should bind to FcgammaRs, while all except IgG6(a) and the putative IgG5 subclass proteins should bind well to porcine FcRn, protein A, and protein G.

Porcine reproductive and respiratory syndrome virus subverts repertoire development by proliferation of germline-encoded B cells of all isotypes bearing hydrophobic heavy chain CDR3.

Isolator piglets infected with porcine reproductive and respiratory syndrome virus (PRRSV), which is related to the lactate dehydrogenase-elevating virus of mice, develop severe hypergammaglobulinemia, lymph node adenopathy, and autoimmune disease. Many of the polyclonally activated B cell clones bear hydrophobic H chain CDR3s (HCDR3s) and are disseminated to most lymphoid tissues. We show in this study that B cells with identical hydrophobic HCDR3s are expressed with all major isotypes in PRRSV-infected piglets (PIPs), explaining why PRRSV-induced hypergammaglobulinemia is seen in all major isotypes. Up to one-third of randomly selected VDJ clones from the respiratory tract of PIPs have hydrophobic HCDR3s exclusively bearing VDJ rearrangements with CDR1, CDR2, and nearly intact DH segments in germline configuration. These HCDR3s are long and D(H)A and D(H)B are exclusively used in reading frame 3. A minimal tripeptide motif containing three hydrophobic amino acids (Leu, Val, and Ile) or any two plus alanine is common to this hydrophobic patch. We propose that PRRSV infection causes generalized Ag-independent B cell activation and hypergammaglobulinemia with biased expansion of a subpopulation of the preimmune repertoire with hydrophobic binding sites that normally disappears during Ag-driven repertoire diversification. Elevated Ig levels in PIP cannot be explained as antiviral Abs; some Igs can account for autoantibodies to dsDNA and Golgi, whereas those with hydrophobic binding sites may account for the Ig aggregates seen in PIPs and lactate dehydrogenase-elevating virus-infected mice. This diversion from normal repertoire development may explain the delayed immune response to PRRSV.

Antibody repertoire development in fetal and neonatal piglets. XIII. Hybrid VH genes and the preimmune repertoire revisited.

The expressed porcine VH genes belong to the VH3 family (clan), four of which, VHA, VHB, VHC, and VHE, alone comprise approximately 80% of the preimmune repertoire. However, so-called "hybrid" VH genes that use CDR1 of one VH gene and the CDR2 of another are frequently encountered. We studied > 3000 cloned VDJs and found that such hybrids can contribute up to 10% of the preimmune repertoire. Based on the 1) recovery of hybrid VH genes from bacterial artificial chromosome clones, 2) frequency of occurrence of certain hybrids in the preimmune repertoire, and 3) failure to recover equal numbers of reciprocal hybrids, we concluded that some chimeric genes are present in the genome and are not PCR artifacts. Two chimeric germline genes (VHZ and VHY), together with VHF and the four genes mentioned above, constitute the major VH genes and these account for > 95% of the preimmune repertoire. Diversification of the preimmune IgG and IgM repertoires after environmental exposure was mainly due to somatic hypermutation of major VH genes with no evidence of gene conversion. Somatic hypermutation was 3- to 10-fold higher in CDRs than in framework regions, most were R mutations and transversions and transitions equally contributed. Data were used to 1) develop an index to quantify the degree of VH repertoire diversification and 2) establish a library of 29 putative porcine VH genes. One-third of these genes are chimeric genes and their sequences suggest that the porcine VH genome developed by duplication and splicing from a small number of prototypic genes.

Antibody repertoire development in fetal and neonatal piglets. XVII. IgG subclass transcription revisited with emphasis on new IgG3.

Fetal piglets offer an in vivo model for determining whether Ag-independent IgG subclass transcription proceeds in a manner that differs from subclass transcription in pigs exposed to environmental Ags and TLR ligands. Our data from approximately 12,000 Cgamma clones from > 60 piglets provide the first report on the relative usage of all known porcine Cgamma genes in fetal and young pigs. Studies revealed that among the six Cgamma genes, allelic variants of IgG1 comprised 50-80% of the repertoire, and IgG2 alleles comprised < 10% in nearly all tissues. However, relative transcription of allelic variants of IgG1 randomly deviate from the 1:1 ratio expected in heterozygotes. Most surprising was the finding that IgG3 accounted for half of all Cgamma transcripts in the ileal Peyer's patches (IPPs) and mesenteric lymph nodes but on average only approximately 5% of the clones from the thymus, tonsil, spleen, peripheral blood, and bone marrow of newborns. Lymphoid tissues from late term fetuses revealed a similar expression pattern. Except for IgG3 in the IPPs and mesenteric lymph nodes, no stochastic pattern of Cgamma expression during development was seen in animals from mid-gestation through 5 mo. The age and tissue dependence of IgG3 transcription paralleled the developmental persistence of the IPP, and its near disappearance corresponds to the diversification of the preimmune VDJ repertoire in young piglets. We hypothesize that long-hinged porcine IgG3 may be important in preadaptive responses to T cell-independent Ags similar to those described for its murine namesake.

Identification of IgF, a hinge-region-containing Ig class, and IgD in Xenopus tropicalis.

Only three Ig isotypes, IgM, IgX, and IgY, were previously known in amphibians. Here, we describe a heavy-chain isotype in Xenopus tropicalis, IgF (encoded by C(phi)), with only two constant region domains. IgF is similar to amphibian IgY in sequence, but the gene contains a hinge exon, making it the earliest example, in evolution, of an Ig isotype with a separately encoded genetic hinge. We also characterized a gene for the heavy chain of IgD, located immediately 3' of C(mu), that shares features with the C(delta) gene in fish and mammals. The latter gene contains eight constant-region-encoding exons and, unlike the chimeric splicing of muC(H)1 onto the IgD heavy chain in teleost fish, it is expressed as a unique IgD heavy chain. The IgH locus of X. tropicalis shows a 5' V(H)-D(H)-J(H)-C(mu)-C(delta)-C(chi)-C(upsilon)-C(phi) 3' organization, suggesting that the mammalian and amphibian Ig heavy-chain loci share a common ancestor.

Development of the neonatal B and T cell repertoire in swine: implications for comparative and veterinary immunology.

Birth in all higher vertebrates is at the center of the critical window of development in which newborns transition from dependence on innate immunity to dependence on their own adaptive immunity, with passive maternal immunity bridging this transition. Therefore we have studied immunological development through fetal and early neonatal life. In swine, B cells appear earlier in fetal development than T cells. B cell development begins in the yolk sac at the 20th day of gestation (DG20), progresses to fetal liver at DG30 and after DG45 continues in bone marrow. The first wave of developing T cells is gammadelta cells expressing a monomorphic Vdelta rearrangement. Thereafter, alphabeta T cells predominate and at birth, at least 19 TRBV subgroups are expressed, 17 of which appear highly homologous with those in humans. In contrast to the T cell repertoire and unlike humans and mice, the porcine pre-immune VH (IGHV-D-J) repertoire is highly restricted, depending primarily on CDR3 for diversity. The V-KAPPA (IGKV-J) repertoire and apparently also the V-LAMBDA (IGLV-J) repertoire, are also restricted. Diversification of the pre-immune B cell repertoire of swine and the ability to respond to both T-dependent and T-independent antigen depends on colonization of the gut after birth in which colonizing bacteria stimulate with Toll-like receptor ligands, especially bacterial DNA. This may explain the link between repertoire diversification and the anatomical location of primary lymphoid tissue like the ileal Peyers patches. Improper development of adaptive immunity can be caused by infectious agents like the porcine reproductive and respiratory syndrome virus that causes immune dysregulation resulting in immunological injury and autoimmunity.