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Odors may be pleasant or unpleasant and in practice, pleasant odors are attractive while unpleasant odors are repellent. However, an odor that is noxious to one species may be attractive to another. Plants, predators, and pathogens may enhance their transmission by manipulating these signals. This may be especially significant when odors attract arthropod disease vectors. Odor detection may also be important in small prey species for evasion of macropredators such as large carnivores. Conversely, pleasant odors may identify family members, parents, or sexual partners. They may also generate signals of good health or fitness and contribute to the process of mate selection. In this review, we seek to integrate these odor-driven processes into a coherent pattern of behaviors that serve to complement the innate and adaptive immune systems. It may be considered the ‘behavioral immune system’.

Antimicrobial peptides (AMPs) are a class of natural peptides with varying numbers of amino acids. They are principal components of innate immunity in vertebrates, encoding natural antibiotics and providing a protective response against a broad range of microbes including those responsible for tuberculosis, an important disease in bison. NK-lysins are AMPs that have been described in various organisms and are coded by a single gene in several mammalian species, including human. Recently, we described a family of 4 NK-lysin genes in cattle. Here, we examined NK-lysin genes in bison and identified 4 bison paralogs (NK1, NK2A, NK2B, and NK2C), although the current bison genome assembly annotates only 2 (NK1 and NK2). Sequence and phylogenetic analysis support the triplication of NK2 prior to the most recent common ancestor of bison and cattle. Comparative mapping of bison and cattle paralogs indicates that the NK-lysin family is located on bison chromosome 11 with well-conserved synteny of flanking genes relative to cattle. The 3 bison NK-lysin2 genes share high sequence similarity with each other. RNA-seq analysis demonstrates that NK2A, NK2B, and NK2C are expressed primarily in the lung, whereas NK1 is expressed at low levels in all tissues studied. This tissue expression pattern differs from that previously reported for cattle, suggesting some divergence in function since the evolutionary separation of the 2 species.

Subclinical illness associated with infection is thought to reduce performance and increase production costs in feedlot cattle, but underlying components remain largely unidentified. Vaccination is frequently used in feedlot settings but producers lack metrics that evaluate the effectiveness of vaccination programs. The goal of this study was to determine if levels of serum neutralizing antibody titers were predictive of levels of vaccine protection in a commercial setting. During this four-year study, Angus-Nellore steers housed in a production feedlot setting were assigned to 1 of 3 vaccine treatments: killed vaccine (kV), modified live virus (MLV) vaccine, or no vaccine (control), and were challenged with a noncytopathic 1b field strain of bovine viral diarrhea virus. Rectal temperature and levels of circulating lymphocytes and platelets were monitored following challenge. While no animals were diagnosed as clinically ill with respiratory disease, indicators of disease (pyrexia, lymphopenia, and thrombocytopenia) were observed. The MLV treatment elicited higher antibody titers to the vaccination than the kV, and calves in the MLV treatment had higher mean titers at challenge. The year that elicited the highest antibody response to the vaccination and the year with the lowest frequency of phenotypic responses to the challenge were not concurrent. The MLV treatment had the highest proportion, 34.68%, of animals that were protected against the challenge regardless of the pre-challenge antibody titer and had the fewest number of lymphopenia cases in response to the challenge. Both vaccine treatments mitigated thrombocytopenia when compared to the control treatment, and the MLV treatment reduced lymphopenia; however, these symptoms were not completely eliminated in vaccinated animals. Pyrexia was present in 40.11% of the animals, but no difference in the frequency of cases between treatments was observed. Pre-challenge vaccination response was not indicative of the level of protection nor was anamnestic antibody response correlated with health status.

In this study, we report an approach to characterize individual BoLA haplotypes using cells from parthenogenetic bovine embryos derived from slaughterhouse ovaries. Eight of the 15 parthenogenetic embryos so obtained had not undergone meiotic recombination on the BoLA region and were suitable to describe BoLA haplotypes. Detailed analysis of the BoLA class IIa region identified seven different class IIa haplotypes, including six not previously described and two new alleles of BoLA-DQA and one BoLA-DQB. Our method provided reliable sources of homozygous DNA to describe BoLA haplotypes.

Appaloosa horses are predisposed to equine recurrent uveitis (ERU), an immune-mediated disease characterized by recurring inflammation of the uveal tract in the eye, which is the leading cause of blindness in horses. Nine genetic markers from the ECA1 region responsible for the spotted coat color of Appaloosa horses, and 13 microsatellites spanning the equine major histocompatibility complex (ELA) on ECA20, were evaluated for association with ERU in a group of 53 Appaloosa ERU cases and 43 healthy Appaloosa controls. Three markers were significantly associated (corrected P-value

The major histocompatibility complex (MHC) in mammals codes for antigen-presenting proteins. For this reason, the MHC is of great importance for immune function and animal health. Previous studies revealed this gene-dense and polymorphic region in river buffalo to be on the short arm of chromosome 2, which is homologous to cattle chromosome 23. Using cattle-derived STS markers and a river buffalo radiation hybrid (RH) panel (BBURH5000 ), we generated a high-resolution RH map of the river buffalo MHC region. The buffalo MHC RH map (cR5000 ) was aligned with the cattle MHC RH map (cR12000 ) to compare gene order. The buffalo MHC had similar organization to the cattle MHC, with class II genes distributed in two segments, class IIa and class IIb. Class IIa was closely associated with the class I and class III regions, and class IIb was a separate cluster. A total of 53 markers were distributed into two linkage groups based on a two-point LOD score threshold of ≥8. The first linkage group included 32 markers from class IIa, class I and class III. The second linkage group included 21 markers from class IIb. Bacterial artificial chromosome clones for seven loci were mapped by fluorescence in situ hybridization on metaphase chromosomes using single- and double-color hybridizations. The order of cytogenetically mapped markers in the region corroborated the physical order of markers obtained from the RH map and served as anchor points to align and orient the linkage groups.

In cattle (Bos taurus), there is evidence of more than 50 alleles of BoLA-DQB (bovine lymphocyte antigen DQB) that are distributed across at least five DQB loci, making this region one of the most complex in the BoLA gene family. In this study, DQB alleles were analysed for the water buffalo (Bubalus bubalis), another economically important bovine species. Twelve alleles for Bubu-DQB (Bubalis bubalis DQB) were determined by nucleotide sequence analysis. A phylogenetic analysis revealed numerous trans-species polymorphisms, with alleles from water buffalo assigned to at least three different loci (BoLA-DQB1, BoLA-DQB3 and BoLA-DQB4) that are also found in cattle. These presumptive loci were analysed for patterns of synonymous (d(S)) and non-synonymous (d(N)) substitution. Like BoLA-DQB1, Bubu-DQB1 was observed to be under strong positive selection for polymorphism. We conclude that water buffalo and cattle share the current arrangement of their DQB region because of their common ancestry.

The assembled genomic sequence of the horse major histocompatibility complex (MHC) (equine lymphocyte antigen, ELA) is very similar to the homologous human HLA, with the notable exception of a large segmental duplication at the boundary of ELA class I and class III that is absent in HLA. The segmental duplication consists of a ∼ 710 kb region of at least 11 repeated blocks: 10 blocks each contain an MHC class I-like sequence and the helicase domain portion of a BAT1-like sequence, and the remaining unit contains the full-length BAT1 gene. Similar genomic features were found in other Perissodactyls, indicating an ancient origin, which is consistent with phylogenetic analyses. Reverse-transcriptase PCR (RT-PCR) of mRNA from peripheral white blood cells of healthy and chronically or acutely infected horses detected transcription from predicted open reading frames in several of the duplicated blocks. This duplication is not present in the sequenced MHCs of most other mammals, although a similar feature at the same relative position is present in the feline MHC (FLA). Striking sequence conservation throughout Perissodactyl evolution is consistent with a functional role for at least some of the genes included within this segmental duplication.

Summary. Genomic DNAs from animals representing six breeds of cattle (Angus, Brahman, Hereford, Holstein, Jersey and Texas Longhorn) were screened with cloned gene probes in a search for restriction fragment length polymorphisms (RFLPs). Eleven RFLPs were identified using seven different probes: growth hormone, prolactin, osteonectin, α A-crystallin, γ crystallin, fibronectin and 21-steroid hydroxylase. The frequencies of the alleles identified by each probe were calculated and compared in a limited sampling of the six bovine breeds. These polymorphisms greatly enhance the pool of immunogenetic, biochemical and molecular markers available in cattle for linkage analysis, testing of parentage, and distinction of breeds.

Mammalian sex chromosomes are the remnants of an ancient autosomal pair present in the ancestral mammalian karyotype. As a consequence of random decay and chromosome rearrangements over evolutionary time, Y chromosome gene repertoires differ between eutherian lineages. To investigate the gene repertoire and transcriptional analysis of the domestic cat Y chromosome, and their potential roles in spermatogenesis, we obtained full-length cDNA sequences for all known Y genes and their X chromosome gametologues and used those sequences to create a BAC-based physical map of the X-degenerate region. Our results indicate the domestic cat Y chromosome has retained most X-degenerate genes that were present on the ancestral eutherian Y chromosome. Transcriptional analysis revealed that most feline X-degenerate genes have retained housekeeping functions shared by their X chromosome partners and have not been specialized for testis-specific functions. Physical mapping data indicate that the cat SRY gene is present as multiple functional copies and that very little of the felid Y chromosome may be single copy. X-Y gene divergence time estimates obtained using Bayesian methods confirm an early origin of Stratum 1 genes prior to the origin of therian mammals. We observed no statistical difference in the ages of Stratum 2 and Stratum 3 gene pairs, suggesting that eutherian and marsupial Stratum 2 genes may have been independently retained in each lineage.

NK-lysin is an antimicrobial peptide and effector protein in the host innate immune system. It is coded by a single gene in humans and most other mammalian species. In this study, we provide evidence for the existence of four NK-lysin genes in a repetitive region on cattle chromosome 11. The NK2A, NK2B, and NK2C genes are tandemly arrayed as three copies in similar to 30-35-kb segments, located 41.8 kb upstream of NK1. All four genes are functional, albeit with differential tissue expression. NK1, NK2A, and NK2B exhibited the highest expression in intestine Peyer's patch, whereas NK2C was expressed almost exclusively in lung. The four peptide products were synthesized ex vivo, and their antimicrobial effects against both Gram-positive and Gram-negative bacteria were confirmed with a bacteria-killing assay. Transmission electron microcopy indicated that bovine NK-lysins exhibited their antimicrobial activities by lytic action in the cell membranes. In summary, the single NK-lysin gene in other mammals has expanded to a four-member gene family by tandem duplications in cattle; all four genes are transcribed, and the synthetic peptides corresponding to the core regions are biologically active and likely contribute to innate immunity in ruminants.

The bovine major histocompatibility complex (MHC) or BoLA is organized differently from typical mammalian MHCs in that a large portion of the class II region, called class IIb, has been transposed to a position near the centromere on bovine chromosome 23. Gene mapping indicated that the rearrangement resulted from a single inversion, but the boundaries and gene content of the inverted segment have not been fully determined. Here, we report the genomic sequence of BoLA IIb. Comparative sequence analysis with the human MHC revealed that the proximal inversion breakpoint occurred approximately 2.5 kb from the 3[prime] end of the glutamate-cysteine ligase, catalytic subunit (GCLC) locus and that the distal breakpoint occurred about 2 kb from the 5[prime] end from a divergent class IIDR[beta]-like sequence designated DSB. Gene content, order and orientation of BoLA IIb are consistent with the single inversion hypothesis when compared with the corresponding region of the human class II MHC (HLA class II). Differences with HLA include the presence of a single histone H2B gene located between the proteasome subunit, beta type, 9 (PSMB9) and DMB loci and a duplicated TAP2 with a variant splice site. BoLA IIb spans approximately 450 kb DNA, with 20 apparently intact genes and no obvious pseudogenes. The region contains 227 simple sequence repeats (SSRs) and approximately 167 kb of retroviral-related repetitive DNA. Nineteen of the 20 genes identified in silico are supported by bovine EST data indicating that the functional gene content of BoLA IIb has not been diminished because it has been transposed from the remainder of BoLA genes.

1 McGowan C. & Reith M. E. (1999) Mol Ecol 8, 1761– 63. 2 Bristol G. & Andersen R. D. (1995) C-primer. Department of Medicine, Hem-Onc and the Department of Biochemistry, School of Medicine, University of California, Los Angeles, USA. 3 Allendorf F. W. & Thorgaard G. H. (1984) Tetraploidy and the evolution of salmonid fishes. In: Evolutionary Genetics of Fishes (Ed. by B.J. Turner), pp. 1–46. Plenum Press, New York, USA. 4 Woram R. A. et al. (2004) Genome 47, 304–15.

The major histocompatibility complex (MHC) in humans is a polymorphic assemblage of over 70 genes which play key roles in the regulation of the immune response (TROWSDALE et al. 1991). The bovine MHC (bovine lymphocyte antigen, BoLA) was assigned to a rather broad region of cattle chromosome 23 (ql3-23) by sequential Q-banding and isotopic in situ hybridization (FRIES et al. 1986; HEDIGER et al. 1991). In the present study, we use sequential RBA-banding and fluorescent in situ hybridization

Development of a dense map of the horse genome is key to efforts aimed at identifying genes controlling health, reproduction, and performance. We herein report a high-resolution gene map of the horse ( Equus caballus ) X chromosome (ECAX) generated by developing and typing 116 gene-specific and 12 short tandem repeat markers on the 5,000-rad horse × hamster whole-genome radiation hybrid panel and mapping 29 gene loci by fluorescence in situ hybridization. The human X chromosome sequence was used as a template to select genes at 1-Mb intervals to develop equine orthologs. Coupled with our previous data, the new map comprises a total of 175 markers (139 genes and 36 short tandem repeats, of which 53 are fluorescence in situ hybridization mapped) distributed on average at ≈880-kb intervals along the chromosome. This is the densest and most uniformly distributed chromosomal map presently available in any mammalian species other than humans and rodents. Comparison of the horse and human X chromosome maps shows remarkable conservation of gene order along the entire span of the chromosomes, including the location of the centromere. An overview of the status of the horse map in relation to mouse, livestock, and companion animal species is also provided. The map will be instrumental for analysis of X linked health and fertility traits in horses by facilitating identification of targeted chromosomal regions for isolation of polymorphic markers, building bacterial artificial chromosome contigs, or sequencing.

Comparative genomics has served as a backbone for the rapid development of gene maps in domesticated animals. The integration of this approach with radiation hybrid (RH) analysis provides one of the most direct ways to obtain physically ordered comparative maps across evolutionarily diverged species. We herein report the development of a detailed RH and comparative map for horse chromosome 17 (ECA17). With markers distributed at an average interval of every 1.4 Mb, the map is currently the most informative among the equine chromosomes. It comprises 75 markers (56 genes and 19 microsatellites), of which 50 gene specific and 5 microsatellite markers were generated in this study and typed to our 5000-rad horse × hamster whole genome RH panel. The markers are dispersed over six RH linkage groups and span 825 cR 5000 . The map is among the most comprehensive whole chromosome comparative maps currently available for domesticated animals. It finely aligns ECA17 to human and mouse homologues (HSA13 and MMU1, 3, 5, 8, and 14, respectively) and homologues in other domesticated animals. Comparisons provide insight into their relative organization and help to identify evolutionarily conserved segments. The new ECA17 map will serve as a template for the development of clusters of BAC contigs in regions containing genes of interest. Sequencing of these regions will help to initiate studies aimed at understanding the molecular mechanisms for various diseases and inherited disorders in horse as well as human.

A low-density, male-based linkage map was constructed as one of the objectives of the International Equine Gene Mapping Workshop. Here we report the second generation map based on testing 503 half-sibling offspring from 13 sire families for 344 informative markers using the CRIMAP program. The multipoint linkage analysis localized 310 markers (90%) with 257 markers being linearly ordered. The map included 34 linkage groups representing all 31 autosomes and spanning 2262 cM with an average interval between loci of 10.1 cM. This map is a milestone in that it is the first map with linkage groups assigned to each of the 31 automosomes and a single linkage group to all but three chromosomes.

Summary Seventy-five individuals of Bubalus bubalis belonging to four different breeds, three of river buffalo and one of swamp buffalo, were studied for polymorphism in MHC DRB (Bubu-DRB) and DRA (Bubu-DRA) loci. Eight alleles of Bubu-DRB were found, and all alleles in the swamp type were shared with the three river breeds. All alleles sampled from the breed of European origin (Mediterranean) were present in breeds sampled in Brazil, thus variability of this locus may have been preserved to a great extent in the more recently founded Brazilian population. Bubu-DRB alleles contained higher proportions of synonymous vs. non-synonymous substitutions in the non-peptide-binding sites (PBS) region, in contrast to the pattern of variation found in BoLA-DRB3, the orthologous locus in cattle. This indicated that either the first domain exon (exon 2) of Bubu-DRB has not undergone as much recombination and/or gene conversion as in cattle alleles, or Bubu-DRB may be more ancient than BoLA-DRB3 alleles. Phylogenetic analysis of DRB alleles from Bubalus, Syncerus c. caffer, the Cape buffalo, and domestic cattle demonstrated transspecies polymorphism. Water buffalo contained two alleles of DRA that differed from each other in two amino acid positions, including one in the PBS (α22) that was also shared with Anoa depressicornis, the anoa. Discovery of variation in DRA was surprising as the first domain of DRA is a highly conserved polypeptide in mammals in general and especially in ruminants, where no other substitution in PBS was seen.

Fluorescence in situ hybridization (FISH) analyses were used to order 16 bacterial artificial chromosomes (BAC) clones containing loci from the bovine lymphocyte antigen (BoLA) class I and III regions of bovine chromosome 23 (BTA23). Fourteen of these BACs were assigned to chromosomal band locations of mitotic and pachytene chromosomes by single- and dual-colour FISH. Dual-colour FISH confirmed that class II DYA is proximal to and separated from BoLA class I genes by approximately three chromosome bands. The FISH results showed that tumour necrosis factor alpha (TNFA), heat shock protein 70 (HSP70.1) and 21 steroid dehydrogenase (CYP21) are closely linked in the region of BTA23 band 22 along with BoLA class I genes, and that male enhanced antigen (MEA) mapped between DYA and the CYP21/TNFA/HSP70.1 gene region. All BAC clones containing BoLA class I genes mapped distal to CYP21/TNFA/HSP70.1 and centromeric to prolactin (PRL). Myelin oligodendrocyte glycoprotein (MOG) was shown to be imbedded within the BoLA class I gene cluster. The cytogenetic data confirmed that the disrupted distribution of BoLA genes is most likely the result of a single large chromosomal inversion. Similar FISH results were obtained when BoLA DYA and class I BAC clones were mapped to discrete chromosomal locations on the BTA homologue in white-tailed deer, suggesting that this chromosomal inversion predates divergence of the advanced ruminant families from a common ancestor.

We report fluorescence in-situ hybridization (FISH) and somatic cell hybrid mapping data for 13 different horse genes (ANP, CD2, CLU, CRISP3, CYP17, FGG, IL1RN, IL10, MMP13, PRM1, PTGS2, TNFA and TP53). Primers for PCR amplification of intronic or untranslated regions were designed from horse-specific DNA or mRNA sequences in GenBank. Two different horse bacterial artificial chromosome (BAC) libraries were screened with PCR for clones containing these 13 Type I loci, nine of which were found in the libraries. BAC clones were used as probes in dual colour FISH to confirm their precise chromosomal origin. The remaining four genes were mapped in a somatic cell hybrid panel. All chromosomal assignments except one were in agreement with human–horse ZOO-FISH data and revealed new and more detailed information on the equine comparative map. CLU was mapped by synteny to ECA2 while human–horse ZOO-FISH data predicted that CLU would be located on ECA9. The assignment of IL1RN permitted analysis of gene order conservation between HSA2 and ECA15, which identified that an event of inversion had occurred during the evolution of these two homologous chromosomes.

The hypothesis that the conservation of sex-chromosome-linked genes among placental mammals could be extended to the horse genome was tested using the UCDavis horse–mouse somatic cell hybrid (SCH) panel. By exploiting the fluorescence in-situ hybridization (FISH) technique to localize an anchor locus, X-inactivation-specific transcript (XIST) on the horse X chromosome, together with the fragmentation and translocation of the X- and Y-chromosome fragments in a somatic cell hybrid panel, we regionally assigned 13 type I and 13 type II (microsatellite) markers to the horse X- and Y-chromosomes. The synteny groups that correspond to horse X- and Y-chromosomes were identified by synteny mapping of sex-specific loci zinc finger protein X-linked (ZFX), zinc finger protein Y-linked (ZFY) and sex-determining region Y (SRY) on the SCH panel. A non-pseudoautosomal gene in the human steroid sulfatase (STS) was identified in both X- and Y-chromosome- containing clones. The regional order of the X-linked type I markers examined in this study, from Xp- to Xq-distal, was [STS- X, the voltage-gated chloride channel 4 (CLCN4)], [ZFX, delta- aminolevulinate synthase 2 (ALAS2)], XIST, coagulation factor IX (F9) and [biglycan (BGN), equine F18, glucose-6-phosphate dehydrogenase (G6PD)] (precise marker order could not be determined for genes within the same brackets). The order of the Y-linked type I markers was STS-Y, SRYand ZFY. These orders are the same arrangements as reported for the human X- and Y-chromosomes, supporting the conservation of genomic organization between the human and the horse sex chromosomes. Regional ordering of X- linked type I and microsatellite markers provides the first integration of type I and type II markers in the horse X chromosome.

Equine combined immunodeficiency disease (CID) is caused by homozygosity for an autosomal recessive gene. To identify linked markers for the disease, we studied a family segregating for the equine CID gene. A stallion and 19 of his CID-affected offspring were tested for marker segregation at 23 microsatellite DNA loci. His CID-affected offspring inherited only one of his two alleles at the HTG8 and HTG4 loci, namely HTG8-186 and HTG4-124, respectively. Lod scores for linkage to the CID gene using a theta of 0.01 were 5.34 for HTG8 and 2.37 for HTG4. The apparent genotypes also suggested linkage disequilibrium between the HTG8-186 allele and the gene for CID. The gene for the DNA protein kinase catalytic subunit (DNA-PK) was recently suggested as a candidate gene for equine CID. A defect of this gene causes a disease in mice that is similar to equine CID. Therefore, we investigated whether this gene might be associated with the microsatellite markers. Analysis of a somatic cell hybrid panel demonstrated synteny of DNA-PK with HTG4 and HTG8 (Kentucky Synteny Group 3). Fluorescence in situ hybridization (FISH) studies demonstrated that DNA-PK is located on horse chromosome ECA9p12. This work supports the hypothesis of DNA-PK as the probable cause of equine CID.

Two genomic clones of the villin (VIL) gene were independently hybridized on river buffalo (Bubalus bubalis, BBU), sheep (Ovis aries, OAR) and goat (Capra hircus, CHI) chromosomes by using sequential fluorescence in situ hybridization (FISH) and R-banding (RBP- and RBA-banding). Clear hybridization signals revealed that VIL is located in BBU 2q33, OAR 2q33 and CHI 2q33. These chromosomes and chromosome bands are believed to be homologous and the VIL locus is the same as that previously found on cattle chromosome 2q43. VIL localization in these three species allows us tentatively to assign all cattle U17 to BBU and CHI 2q and to extend the physical map to OAR 2q.

Genes within the major histocompatibility complex (MHC) encode proteins involved in innate and adaptive immune responses. Genetic variation in this region can influence the immune response of an individual animal to challenges from a variety of pathogens; however, a complete documentation of genetic variation in the MHC is lacking for most domestic animals, including horses. To provide additional genetic markers for study of the horse MHC, or ELA (equine lymphocyte antigen), we identified 37 polymorphic microsatellite repeats in ELA and used these variations separately and together with published SNPs to investigate linkage disequilibrium (LD) and haplotype structure in a sample of Thoroughbred horses. ELA SNPs alone detected little LD, but microsatellites, either separately or combined with SNPs, revealed substantially more LD. A subset of markers in very high LD across the breadth of ELA may be predictive of structural polymorphisms or linked epistases that are important drivers of haplotype structure in Thoroughbreds.

Mutations in the mouse microphthalmia (mi) gene affect the development of a number of cell types including melanocytes, osteoclasts and mast cells. Recently, mutations in the human mi gene (MITF) were found in patients with Waardenburg Syndrome type 2 (WS2), a dominantly inherited syndrome associated with hearing loss and pigmentary disturbances. We have characterized the molecular defects associated with eight murine mi mutations, which vary in both their mode of inheritance and in the cell types they affect. These molecular data, combined with the extensive body of genetic data accumulated for murine mi, shed light on the phenotypic and developmental consequences of mi mutations and offer a mouse model for WS2.

Outputs from a physiologically based toxicokinetic (PB-TK) model for fish were visualized by mapping time-series data for specific tissues onto a three-dimensional representation of a rainbow trout. The trout representation was generated in stepwise fashion: 1) cross-section images were obtained from an anesthetized fish using a magnetic resonance imaging system, 2) images were processed to classify tissue types and eliminate unnecessary detail. 3) processed images were imported to a visualization software package (Application Visualization System) to create a three-dimensional representation of the fish, encapsulating five volumes corresponding to the liver, kidney, muscle, gastrointestinal tract, and fat, Kinetic data for the disposition of pentachloroethane in trout were generated using a PB-TK model. Model outputs were mapped onto corresponding tissues volumes, representing chemical concentration as color intensity. The workstation software was then used to animate the images, illustration the accumulation of pentachloroethane in each tissue during a continuous branchial (gill) exposure. Images p952-a Figure 1. a Figure 1. b Figure 2. Figure 3. Figure 4. Figure 5. Figure 6. Figure 7. Figure 8. a Figure 8. b

A bovine genomic library was screened with a 2.3 kilobase (kb) human HSP70 cDNA probe. A recombinant phage containing one complete HSP70 gene and a portion of a second HSP70 gene was isolated. Both HSP70 coding regions showed high DNA and deduced amino acid sequence homology, chromosomal location, and gene arrangement; we conclude that these sequences are from the bovine homologues of human HSP70‐1 and ‐2.

Historical hybridization between Bison bison (bison) and Bos taurus (cattle) has been well documented and resulted in cattle mitochondrial DNA (mtDNA) introgression, previously identified in six different bison populations. In order to examine Y chromosome introgression, a microsatellite marker (BYM-1) with non-overlapping allele size distributions in bison and cattle was isolated from a bacterial artificial chromosome (BAC) clone, and was physically assigned to the Y chromosome by fluorescence in situ hybridization. BYM-1 genotypes for a sample of 143 male bison from 10 populations, including all six populations where cattle mtDNA haplotypes were previously identified, indicated that cattle Y chromosome introgression had not occurred in these bison populations. The differential permeability of uniparentally inherited markers to introgression is consistent with observations of sterility among first generation hybrid males and a sexual asymmetry in the direction of hybridization favouring matings between male bison and female cattle.

To explore further the genetics of susceptibility to skin tumor promotion in inbred mice, several aspects of responsiveness to 12-O-tetradecanoylphorbol-13-acetate (TPA) were examined in C3H/He mice and segregating crosses between this mouse strain and C57BL/6 mice as well as BXD and BXH recombinant inbred (RI) strains. Dose-response relationships were established for skin tumor promotion by TPA following initiation with 7,12-dimethylbenz[a]anthracene in C3H/He and B6C3F1, as well as several other mouse stocks and strains included for comparison. The relative responsiveness to TPA skin tumor promotion was: SENCAR much greater than DBA/2 greater than C3H/He approximately B6D2F1 greater than B6C3F1 much greater than C57BL/6. Analyses of the susceptibility of B6C3F2 and B6C3F1 x C57BL/6 backcross mice suggested that a minimum of two dominant genetic loci control responsiveness to phorbol ester promotion in these mice. Further analysis of BXH and BXD RI strains suggested the presence of four distinct promotion-responsive phenotypes controlled by a minimum of two genetic loci. The existence of a 'hyper-responsive' phenotype in the sets of RI strains, however, suggests that a third, recessive locus also may play a role in controlling responsiveness to TPA promotion. At 48 h after the last of four applications of TPA, marked hyperplasia and an increase in dark basal keratinocytes were observed in C3H/He mice, whereas in B6C3F1 mice the response in these parameters was intermediate between C3H/He and C57BL/6 mice. A marked dermal inflammation, as determined by infiltration of polymorphonuclear cells, was observed in C3H/He and B6C3F1 mice, whereas little was noted in C57BL/6 mice. Furthermore, histological evaluations of selected BXD RI strains revealed a significant correlation between the magnitude of the hyperplasia response and the percentage of mice bearing tumors. The present data, in conjunction with our previous studies, confirm that the major gene(s) controlling susceptibility to tumor promoter induced by TPA in two sensitive strains (i.e. DBA/2 C3H/He) are similar or closely linked to those for induction of sustained hyperplasia. In addition, the present data provide new evidence for a model where allelic differences at a minimum of three loci contribute to gene differences in susceptibility to phorbol ester promotion DBA/2 and C3H/He versus C57BL/6 mice.

Background The cattle MHC is termed the bovine leukocyte antigen (BoLA) and, along with the MHCs of other ruminants, is unique in its genomic organization. Consequently, correct and reliable gene maps and sequence information are critical to the study of the BoLA region. The bovine genome sequencing project has produced two assemblies (Btau_3.1 and 4.0) that differ substantially from each other and from conventional gene maps in the BoLA region. To independently compare the accuracies of the different sequence assemblies, we have generated a high resolution map of BoLA using a 12,000rad radiation hybrid panel. Seventy-seven unique sequence tagged site (STS) markers chosen at approximately 50 kb intervals from the Btau 2.0 assembly and spanning the IIa-III-I and IIb regions of the bovine MHC were mapped on a 12,000rad bovine radiation hybrid (RH) panel to evaluate the different assemblies of the bovine genome sequence. Results Analysis of the data generated a high resolution RH map of BoLA that was significantly different from the Btau_3.1 assembly of the bovine genome but in good agreement with the Btau_4.0 assembly. Of the few discordancies between the RH map and Btau_4.0, most could be attributed to closely spaced markers that could not be precisely ordered in the RH panel. One probable incorrectly-assembled sequence and three missing sequences were noted in the Btau_4.0 assembly. The RH map of BoLA is also highly concordant with the sequence-based map of HLA (NCBI build 36) when reordered to account for the ancestral inversion in the ruminant MHC. Conclusion These results strongly suggest that studies using Btau_3.1 for analyses of the BoLA region should be reevaluated in light of the Btau_4.0 assembly and indicate that additional research is needed to produce a complete assembly of the BoLA genomic sequences.

To understand the biology and evolution of ruminants, the cattle genome was sequenced to about sevenfold coverage. The cattle genome contains a minimum of 22,000 genes, with a core set of 14,345 orthologs shared among seven mammalian species of which 1217 are absent or undetected in noneutherian (marsupial or monotreme) genomes. Cattle-specific evolutionary breakpoint regions in chromosomes have a higher density of segmental duplications, enrichment of repetitive elements, and species-specific variations in genes associated with lactation and immune responsiveness. Genes involved in metabolism are generally highly conserved, although five metabolic genes are deleted or extensively diverged from their human orthologs. The cattle genome sequence thus provides a resource for understanding mammalian evolution and accelerating livestock genetic improvement for milk and meat production. Fil: Bovine Genome Sequencing and Analysis Consortium. Bovine Genome Sequencing And Analysis Consortium; Estados Unidos Fil: Amadio, Ariel Fernando. Instituto Nacional de Tecnología Agropecuaria. Centro Regional Santa Fe. Estación Experimental Agropecuaria Rafaela; Argentina Fil: Poli, Mario Andres. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigación en Ciencias Veterinarias y Agronómicas. Instituto de Genética; Argentina

A Horse Is a Horse, of Course The history of horse domestication is closely tied to the history of the human society. Wade et al. (p. 865 ) report on the sequencing and provide a single nucleotide polymorphism map of the horse ( Equus caballus ) genome. Horses are a member of the order perissodactyla (odd-toed animals with hooves). The analysis reveals an evolutionarily new centromere on equine chromosome 11 that displays properties of an immature but fully functioning centromere and is devoid of centromeric satellite sequence. The findings clarify the nature of genetic diversity within and across horse breeds and suggest that the horse was domesticated from a relatively large number of females, but few males.

Mammalian sex chromosomes are the remnants of an ancient autosomal pair present in the ancestral mammalian karyotype. As a consequence of random decay and chromosome rearrangements over evolutionary time, Y chromosome gene repertoires differ between eutherian lineages. To investigate the gene repertoire and transcriptional analysis of the domestic cat Y chromosome, and their potential roles in spermatogenesis, we obtained full-length cDNA sequences for all known Y genes and their X chromosome gametologues and used those sequences to create a BAC-based physical map of the X-degenerate region. Our results indicate the domestic cat Y chromosome has retained most X-degenerate genes that were present on the ancestral eutherian Y chromosome. Transcriptional analysis revealed that most feline X-degenerate genes have retained housekeeping functions shared by their X chromosome partners and have not been specialized for testis-specific functions. Physical mapping data indicate that the cat SRY gene is present as multiple functional copies and that very little of the felid Y chromosome may be single copy. X-Y gene divergence time estimates obtained using Bayesian methods confirm an early origin of Stratum 1 genes prior to the origin of therian mammals. We observed no statistical difference in the ages of Stratum 2 and Stratum 3 gene pairs, suggesting that eutherian and marsupial Stratum 2 genes may have been independently retained in each lineage.

A comprehensive second-generation whole genome radiation hybrid (RH II), cytogenetic and comparative map of the horse genome (2n = 64) has been developed using the 5000rad horse × hamster radiation hybrid panel and fluorescence in situ hybridization (FISH). The map contains 4,103 markers (3,816 RH; 1,144 FISH) assigned to all 31 pairs of autosomes and the X chromosome. The RH maps of individual chromosomes are anchored and oriented using 857 cytogenetic markers. The overall resolution of the map is one marker per 775 kilobase pairs (kb), which represents a more than five-fold improvement over the first-generation map. The RH II incorporates 920 markers shared jointly with the two recently reported meiotic maps. Consequently the two maps were aligned with the RH II maps of individual autosomes and the X chromosome. Additionally, a comparative map of the horse genome was generated by connecting 1,904 loci on the horse map with genome sequences available for eight diverse vertebrates to highlight regions of evolutionarily conserved syntenies, linkages, and chromosomal breakpoints. The integrated map thus obtained presents the most comprehensive information on the physical and comparative organization of the equine genome and will assist future assemblies of whole genome BAC fingerprint maps and the genome sequence. It will also serve as a tool to identify genes governing health, disease and performance traits in horses and assist us in understanding the evolution of the equine genome in relation to other species.

A comparative approach that utilizes information from more densely mapped or sequenced genomes is a proven and efficient means to increase our knowledge of the structure of the horse genome. Human chromosome 2 (HSA2), the second largest human chromosome, comprising 243 Mb, and containing 1246 known genes, corresponds to all or parts of three equine chromosomes. This report describes the assignment of 140 new markers (78 genes and 62 microsatellites) to the equine radiation hybrid (RH) map, and the anchoring of 24 of these markers to horse chromosomes by FISH. The updated equine RH maps for ECA6p, ECA15, and ECA18 resulting from this work have one, two, and three RH linkage groups, respectively, per chromosome/chromosome-arm. These maps have a three-fold increase in the number of mapped markers compared to previous maps of these chromosomes, and an increase in the average marker density to one marker per 1.3 Mb. Comparative maps of ECA6p, ECA15, and ECA18 with human, chimpanzee, dog, mouse, rat, and chicken genomes reveal blocks of conserved synteny across mammals and vertebrates.

The genome organizations of eight phylogenetically distinct species from five mammalian orders were compared in order to address fundamental questions relating to mammalian chromosomal evolution. Rates of chromosome evolution within mammalian orders were found to increase since the Cretaceous-Tertiary boundary. Nearly 20% of chromosome breakpoint regions were reused during mammalian evolution; these reuse sites are also enriched for centromeres. Analysis of gene content in and around evolutionary breakpoint regions revealed increased gene density relative to the genome-wide average. We found that segmental duplications populate the majority of primate-specific breakpoints and often flank inverted chromosome segments, implicating their role in chromosomal rearrangement.

Childwick Trust supported J.E.S and M.M.B. Awards from the Morris Animal Foundation and the United States Department of Agriculture National Research Initiate Competitive Grants (JRM, LJA, LCS, BPC), the Texas Higher Education Board (ARP 010366-0162-2001; BPC), Link Endowment (BPC, LCS), and the Dorothy Russell Havemeyer Foundation (BPC). We also wish to thank Ms Louise Pettit for expert technical assistance.

High-resolution gene maps of individual equine chromosomes are essential to identify genes governing traits of economic importance in the horse. In pursuit of this goal we herein report the generation of a dense map of horse chromosome 22 (ECA22) comprising 83 markers, of which 52 represent specific genes and 31 are microsatellites. The map spans 831 cR over an estimated 64 Mb of physical length of the chromosome, thus providing markers at approximately 770 kb or 10 cR intervals. Overall, the resolution of the map is to date the densest in the horse and is the highest for any of the domesticated animal species for which annotated sequence data are not yet available. Comparative analysis showed that ECA22 shares remarkable conservation of gene order along the entire length of dog chromosome 24, something not yet found for an autosome in evolutionarily diverged species. Comparison with human, mouse, and rat homologues shows that ECA22 can be traced as two conserved linkage blocks, each related to individual arms of the human homologue-HSA20. Extending the comparison to the chicken genome showed that one of the ECA22 blocks that corresponds to HSA20q shares synteny conservation with chicken chromosome 20, suggesting the segment to be ancestral in mammals and birds.

Comparative biochemical and histopathological data suggest that a deficiency in the glycogen branching enzyme (GBE) is responsible for a fatal neonatal disease in Quarter Horse foals that closely resembles human glycogen storage disease type IV (GSD IV). Identification of DNA markers closely linked to the equine GBE1 gene would assist us in determining whether a mutation in this gene leads to the GSD IV-like condition. FISH using BAC clones as probes assigned the equine GBE1 gene to a marker deficient region of ECA26q12→q13. Four other genes, ROBO2, ROBO1, POU1F1, and HTR1F, that flank GBE1 within a 10-Mb segment of HSA3p12→p11, were tightly linked to equine GBE1 when analyzed on the Texas A&M University 5000 rad equine radiation hybrid panel, while the GLB1, MITF, RYBP, and PROS1 genes that flank this 10-Mb interval were not linked with markers in the GBE1 group. A polymorphic microsatellite (GBEms1) in a GBE1 BAC clone was then identified and genetically mapped to ECA26 on the Animal Health Trust full-sibling equine reference family. All Quarter Horse foals affected with GSD IV were homozygous for an allele of GBEms1, as well as an allele of the most closely linked microsatellite marker, while a control horse population showed significant allelic variation with these markers. This data provides strong molecular genetic support for the candidacy of the GBE1 locus in equine GSD IV.

A physical map of ordered bacterial artificial chromosome (BAC) clones was constructed to determine the genetic organization of the horse major histocompatibility complex. Human, cattle, pig, mouse, and rat MHC gene sequences were compared to identify highly conserved regions which served as source templates for the design of overgo primers. Thirty-five overgo probes were designed from 24 genes and used for hybridization screening of the equine USDA CHORI 241 BAC library. Two hundred thirty-eight BAC clones were assembled into two contigs spanning the horse MHC region. The first contig contains the MHC class II region and was reduced to a minimum tiling path of nine BAC clones that span approximately 800 kb and contain at least 20 genes. A minimum tiling path of a second contig containing the class III/I region is comprised of 14 BAC clones that span approximately 1.6 Mb and contain at least 34 genes. Fluorescence in situ hybridization (FISH) using representative clones from each of the three regions of the MHC localized the contigs onto ECA20q21 and oriented the regions relative to one another and the centromere. Dual-colored FISH revealed that the class I region is proximal to the centromere, the class II region is distal, and the class III region is located between class I and II. These data indicate that the equine MHC is a single gene-dense region similar in structure and organization to the human MHC and is not disrupted as in ruminants and pigs.

Five genomic clones representing three HSP70 genes of cattle were biotin labeled and independently hybridized to cattle chromosomes. Fluorescence in situ hybridization localized HSP70-2 to Chromosome (Chr) 23 band 22 (the BoLA region), HSP70-3 to Chr 10 band 34, and HSP70-4 to Chr 3 band 13. Since HSP70-1, a fourth HSP70 gene, is tightly linked with HSP70-2 and the BOLA, HSP70-1 was also localized to Chr 23 band 22. The localization of HSP70-4 is the first assignment of a cattle U6 marker; thus, this entire syntenic group is tentatively placed in cattle Chr 3.

A 5000(rad) whole-genome radiation hybrid (RH) panel was created for the horse. The usefulness of the panel for generating physically ordered maps of individual equine chromosomes was tested by typing 24 markers on horse Chromosome 11 (ECA11). The overall retention of markers on this chromosome was 43.6%. Almost complete retention of two of the typed markers--- CA062 and AHT44---clearly indicated the location of thymidine kinase gene on the short arm of ECA11. Seven of the typed markers were FISH mapped to align the RH and cytogenetic maps. With the RH-MAPPER approach, a physically ordered map comprising four linkage groups and incorporating all the markers was obtained. The study provides the first comprehensive map for a horse chromosome that integrates all available mapping data and adds new information that spans the entire length of the equine chromosome. The map clearly underlines the resolving power and utility of the panel and emphasizes the need to have uniformly distributed cytogenetic markers for appropriate alignment of RH map with the chromosome. A comparative status of the ECA11 map in relation to the corresponding human/mouse chromosome is presented.

A new physical map of the bovine genome has been constructed by integrating data from genetic and radiation hybrid maps, and a new bovine BAC map, with the bovine genome draft assembly., Background Cattle are important agriculturally and relevant as a model organism. Previously described genetic and radiation hybrid (RH) maps of the bovine genome have been used to identify genomic regions and genes affecting specific traits. Application of these maps to identify influential genetic polymorphisms will be enhanced by integration with each other and with bacterial artificial chromosome (BAC) libraries. The BAC libraries and clone maps are essential for the hybrid clone-by-clone/whole-genome shotgun sequencing approach taken by the bovine genome sequencing project. Results A bovine BAC map was constructed with HindIII restriction digest fragments of 290,797 BAC clones from animals of three different breeds. Comparative mapping of 422,522 BAC end sequences assisted with BAC map ordering and assembly. Genotypes and pedigree from two genetic maps and marker scores from three whole-genome RH panels were consolidated on a 17,254-marker composite map. Sequence similarity allowed integrating the BAC and composite maps with the bovine draft assembly (Btau3.1), establishing a comprehensive resource describing the bovine genome. Agreement between the marker and BAC maps and the draft assembly is high, although discrepancies exist. The composite and BAC maps are more similar than either is to the draft assembly. Conclusion Further refinement of the maps and greater integration into the genome assembly process may contribute to a high quality assembly. The maps provide resources to associate phenotypic variation with underlying genomic variation, and are crucial resources for understanding the biology underpinning this important ruminant species so closely associated with humans.

The class II loci of the major histocompatibility complex (MHC)of humans (HLA) and mice (H-2) are embedded among other lociof the MHC, which spans approximately 4 mb of DNA and has arecombination length of about 2 cM (Campbell and Trowsdale1993; Himmelbauer et al. 1993).In contrast, recombination analysis of class II genes in thebovine lymphocyte antigen complex (BoLA) revealed two distinctclusters of class II loci about 17 cM apart (Anderson et al. 1988).One cluster designated IIa contains the bovine homologs of thehuman DQ and DR gene families and is tightly linked to otherBoLA genes, including HSP70, CYP21, and several class I loci(Bishop et al. 1994). The second cluster, IIb, contains the DYA,DYB, DOB loci and a novel class II gene designated DIB (Stoneand Muggli-Cockett 1990). The disruption of the BoLA class IIloci into two clusters suggests that the BoLA complex has under-gone significant rearrangements not yet observed in the MHCs ofhumans or mice, but that apparently have occurred in the MHCs ofchickens (Miller et al. 1994) and swine (Smith et al. 1995). Alter-natively, the bovine class IIa and IIb gene clusters could be inphysical proximity but undergo frequent recombinant owing to oneor more recombination hotspots. Recombination hotspots havebeen reported in the HLA and H-2 regions (Shiroishi et al. 1990;Steinmetz et al. 1986; Yoshino et al. 1994), and anomalous re-combination frequencies in this region among individual bullshave been reported (Park et al. 1995).To test whether the BoLA IIb cluster mapped to a differentchromosomal location than the other BoLA loci, we performedfluorescence in situ hybridization (FISH) analysis of bovine Chro-mosome (Chr) 23, using large insert clones containing DYA orclass I sequences as fluorescent probes. In this report, we demon-strate that DNA probes containing bovine DYA sequences map toa different region of Chr 23 than does DNA containing class Isequences, consistent with the reported recombination distance of17 cM between these genes.Recombinant clones of bacterial artificial chromosomes(BACs) containing DYA or class I sequences were isolated byPCR screening of a bovine BAC library (Cai et al. 1995) withprimer pairs that have been previously described (Van Eijk et al.1992; Garber et al. 1994). Authenticity of each clone was con-firmed by sequencing of the respective PCR products and com-paring with published sequences.BAC clones 6 and 76 were identified by PCR as containingDYA and class I sequences, respectively, and DNAs from theseclones were prepared for FISH. The sequence of the PCR productfrom BAC 6 differed at only a single position