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Goats (Capra hircus) are an economically important livestock species providing meat and milk across the globe. They are of particular importance in tropical agri-systems contributing to sustainable agriculture, alleviation of poverty, social cohesion and utilisation of marginal grazing. There are excellent genetic and genomic resources available for goats, including a highly contiguous reference genome (ARS1). However, gene expression information is limited in comparison to other ruminants. To support functional annotation of the genome and comparative transcriptomics we created a mini-atlas of gene expression for the domestic goat. RNA-Seq analysis of 22 transcriptionally rich tissues and cell-types detected the majority (90%) of predicted protein-coding transcripts and assigned informative gene names to more than 1000 previously unannotated protein-coding genes in the current reference genome for goat (ARS1). Using network-based cluster analysis we grouped genes according to their expression patterns and assigned those groups of co-expressed genes to specific cell populations or pathways. We describe clusters of genes expressed in the gastro-intestinal tract and provide the expression profiles across tissues of a subset of genes associated with functional traits. Comparative analysis of the goat atlas with the larger sheep gene expression atlas dataset revealed transcriptional differences between the two species in macrophage-associated signatures. The goat transcriptomic resource complements the large gene expression dataset we have generated for sheep and contributes to the available genomic resources for interpretation of the relationship between genotype and phenotype in small ruminants.

This project sought to investigate the domestic caprid rumen virome by developing a robust viral DNA isolation and enrichment protocol (utilizing membrane filtration, ultra-centrifugation, overnight PEG treatment and nuclease treatment) and using RSD-PCR and high throughput sequencing (HTS) techniques. 3.53% of the reads obtained were analogous to those of viruses denoting Siphoviridae, Myoviridae, Podoviridae, Mimiviridae, Microviridae, Poxviridae, Tectiviridae and Marseillevirus. Most of the sequenced reads from the rumen were similar to those of phages, which are critical in maintaining the rumen microbial populations under its carrying capacity. Though identified in the rumen, most of these viruses have been reported in other environments as well. Improvements in the viral DNA enrichment and isolation protocol are required to obtain data that are more representative of the rumen virome. The 102,130 unknown reads (92.31%) for the goat and 36,241 unknown reads (93.86%) for the sheep obtained may represent novel genomes that need further study.

African bovine trypanosomiasis caused by Trypanosoma sp., is a major constraint on cattle productivity in sub-Saharan Africa. Some African Bos taurus breeds are highly tolerant of infection, but the potentially more productive Bos indicus zebu breeds are much more susceptible. Zebu cattle are well adapted for plowing and haulage, and increasing their tolerance of trypanosomiasis could have a major impact on crop cultivation as well as dairy and beef production. We used three strategies to obtain short lists of candidate genes within QTL that were previously shown to regulate response to infection. We analyzed the transcriptomes of trypanotolerant N'Dama and susceptible Boran cattle after infection with Trypanosoma congolense. We sequenced EST libraries from these two breeds to identify polymorphisms that might underlie previously identified quantitative trait loci (QTL), and we assessed QTL regions and candidate loci for evidence of selective sweeps. The scan of the EST sequences identified a previously undescribed polymorphism in ARHGAP15 in the Bta2 trypanotolerance QTL. The polymorphism affects gene function in vitro and could contribute to the observed differences in expression of the MAPK pathway in vivo. The expression data showed that TLR and MAPK pathways responded to infection, and the former contained TICAM1, which is within a QTL on Bta7. Genetic analyses showed that selective sweeps had occurred at TICAM1 and ARHGAP15 loci in African taurine cattle, making them strong candidates for the genes underlying the QTL. Candidate QTL genes were identified in other QTL by their expression profile and the pathways in which they participate.

To examine differences in cytokine profiles that may confer tolerance/susceptibility to bovine African trypanosomiasis, N'Dama (trypanotolerant, n = 8) and Boran (trypanosusceptible, n = 8) cattle were experimentally challenged with Trypanosoma congolense. Blood samples were collected over a 34-day period, and RNA was extracted from peripheral blood mononuclear cells. The expression levels of a panel of 14 cytokines were profiled over the time course of infection and between breeds. Messenger RNA (mRNA) transcript levels for the IL2, IL8, and IL1RN genes were significantly downregulated across the time course of infection in both breeds. There was an early increase in transcripts for genes encoding proinflammatory mediators ( IFNG, IL1A, TNF, and IL12) in N'Dama by 14 days postinfection (dpi) compared with preinfection levels that was not detected in the susceptible Boran breed. By the time of peak parasitemia, a type 2 helper T cells (TH2)-like cytokine environment was prevalent that was particularly evident in the Boran. Increases in transcripts for the IL6 (29 and 34 dpi) and IL10 (21, 25, and 29 dpi) genes were detected that were higher in the Boran compared with N'Dama. These findings highlight the implications for using murine models to study the bovine immune response to trypanosomiasis, where in some cases cytokine expression patterns differ. Overall, these data suggest that the trypanotolerant N'Dama are more capable of responding very early in infection with proinflammatory and TH1 type cytokines than the trypanosusceptible Boran and may explain why N'Dama control parasitemia more efficiently than Boran during the early stages of infection.

Trypanosomosis, or sleeping sickness, is a major disease constraint on livestock productivity in sub-Saharan Africa. To identify quantitative trait loci (QTL) controlling resistance to trypanosomosis in cattle, an experimental cross was made between trypanotolerant African N'Dama ( Bos taurus ) and trypanosusceptible improved Kenya Boran ( Bos indicus ) cattle. Sixteen phenotypic traits were defined describing anemia, body weight, and parasitemia. One hundred seventy-seven F 2 animals and their parents and grandparents were genotyped at 477 molecular marker loci covering all 29 cattle autosomes. Total genome coverage was 82%. Putative QTL were mapped to 18 autosomes at a genomewise false discovery rate of 2 cross between N'Dama and Boran cattle could produce a synthetic breed with higher trypanotolerance levels than currently exist in the parental breeds.

Chediak-Higashi Syndrome (CHS) is an autosomal recessive disorder that affects several species including mice, humans, and cattle. Evidence based on clinical characteristics and somatic cell genetics suggests that mutations in a common gene cause CHS in the three species. The CHS locus on human chromosome 1 and mouse chromosome 13 encodes a lysosomal trafficking regulator formerly known as LYST, now known as CHS1, and is defective in CHS patients and beige mice, respectively. We have mapped the CHS locus to the proximal region of bovine chromosome 28 by linkage analysis using microsatellite markers previously mapped to this chromosome. Furthermore, we have identified a missense A:T-->G:C mutation that results in replacement of a histidine with an arginine residue at codon 2015 of the CHS1 gene. This mutation is the most likely cause of CHS in Wagyu cattle. In addition, we describe quick, inexpensive, PCR based tests that will permit elimination of the CHS mutation from Wagyu breeding herds.

We have identified substitutions in the 3' untranslated region of the bovine myoglobin gene, one of which affects an MboII restriction enzyme site resulting in a bi-allelic restriction fragment length polymorphism. Co-dominant inheritance of the alleles in three reference families was observed using a polymerase chain reaction--restriction fragment length polymorphism assay. The distribution of the alleles seems characteristic of cattle type--one of the alleles was not detected in purely taurine breeds. Furthermore, we mapped, using the polymerase chain reaction on a bovine-rodent somatic cell hybrid panel, the myoglobin gene to bovine chromosome five. It is therefore syntenic with gamma-interferon and insulin-like growth factor in which we have not found polymorphism. The myoglobin locus therefore serves as a type one marker on bovine chromosome five.

Background African animal trypanosomiasis (AAT) caused by tsetse fly-transmitted protozoa of the genus Trypanosoma is a major constraint on livestock and agricultural production in Africa and is among the top ten global cattle diseases impacting on the poor. Here we show that a functional genomics approach can be used to identify temporal changes in host peripheral blood mononuclear cell (PBMC) gene expression due to disease progression. We also show that major gene expression differences exist between cattle from trypanotolerant and trypanosusceptible breeds. Using bovine long oligonucleotide microarrays and real time quantitative reverse transcription PCR (qRT-PCR) validation we analysed PBMC gene expression in naïve trypanotolerant and trypanosusceptible cattle experimentally challenged with Trypanosoma congolense across a 34-day infection time course. Results Trypanotolerant N'Dama cattle displayed a rapid and distinct transcriptional response to infection, with a ten-fold higher number of genes differentially expressed at day 14 post-infection compared to trypanosusceptible Boran cattle. These analyses identified coordinated temporal gene expression changes for both breeds in response to trypanosome infection. In addition, a panel of genes were identified that showed pronounced differences in gene expression between the two breeds, which may underlie the phenomena of trypanotolerance and trypanosusceptibility. Gene ontology (GO) analysis demonstrate that the products of these genes may contribute to increased mitochondrial mRNA translational efficiency, a more pronounced B cell response, an elevated activation status and a heightened response to stress in trypanotolerant cattle. Conclusion This study has revealed an extensive and diverse range of cellular processes that are altered temporally in response to trypanosome infection in African cattle. Results indicate that the trypanotolerant N'Dama cattle respond more rapidly and with a greater magnitude to infection compared to the trypanosusceptible Boran cattle. Specifically, a subset of the genes analyzed by real time qRT-PCR, which display significant breed differences, could collectively contribute to the trypanotolerance trait in N'Dama.

An international multidisciplinary consortium is conducting a programme of research on the host response to trypanosome infection. This builds upon quantitative trait loci (QTL) mapping which identified genome regions influencing susceptibility to pathology following T. congolense infection in both cattle and mice. The approach uses expression analysis to examine the response of both susceptible and resistant strains and a series of novel informatics tools to identify pathways which are activated as a result of challenge, and those which are differentially used by resistant and susceptible strains. Of particular interest are those pathways which simultaneously satisfy both criteria, i.e. are significantly differentially activated and contain genes within QTL regions. However, it is important to stress that it is not required that the genes within the QTL region are differentially expressed themselves.

High-throughputtechnologies inevitably produce vast quantities of data. This presents challenges in terms of developing effective analysis methods, particularly where the analysis involves combining data derived from different experimental technologies. In this investigation, a systematic approach was applied to combine microarray gene expression data, quantitative trait loci (QTL) data and pathway analysis resources in order to identify functional candidate genes underlying tolerance to Trypanosoma congolense infection in cattle. We automated much of the analysis using Taverna workflows previously developed for the study of trypanotolerance in the mouse model. Pathways represented by genes within the QTL regions were identified, and this list was subsequently ranked according to which pathways were over-represented in the set of genes that were differentially expressed (over time or between tolerant N'dama and susceptible Boran breeds) at various timepoints after T. congolense infection. The genes within the QTLthat played a role in the highest ranked pathways were flagged as good targets for further investigation and experimental confirmation.

Thymic-derived lymphocytes (T cells) recognize and respond to antigens through the mediation of the T-cell receptor complex. Here we report polymorphism at each of the five loci that encode the different components of the T-cell receptor complex in cattle. These genes were mapped on the bovine genome by genetic linkage analysis in the International Bovine Reference Panel (IBRP). These mapping data provide additional type I markers for linking the bovine genetic map with the human and mouse maps and also permit investigation of the effect of T-cell receptor polymorphism on immune responsiveness and disease susceptibility.

Enzymes that lengthen the carbon chain of polyunsaturated fatty acids are key to the biosynthesis of the highly unsaturated fatty acids, arachidonic, eicosapentaenoic and docosahexaenoic acids from linoleic and alpha-linolenic acids. A Mortierella alpina cDNA polyunsaturated fatty acid elongase sequence identified mammalian, amphibian, zebrafish and insect expressed sequence tags (ESTs) in GenBank. Consensus primers were designed in conserved motifs and used to isolate full length cDNA from livers of several fish species by Rapid Amplification of cDNA Ends (RACE). The amplified cDNAs encoded putative open reading frames (ORFs) of 288-294 amino acids that were highly conserved among the fish species. Heterologous expression in yeast, Saccharomyces cerevisiae, demonstrated that all of the ORFs encoded elongases with the ability to lengthen polyunsaturated fatty acid substrates with chain lengths from C18 to C22 and also monounsaturated fatty acids, but not saturated fatty acids. There were differences in the functional competence of the elongases from different fish species. Most of the fish elongases showed a pattern of activity towards different fatty acid substrates in the rank order C18>C20>C22, although the tilapia and turbot elongases had similar activity towards 18:4n-3 and 20:5n-3. The fish elongases generally showed greater activity or similar activities with n-3 than with n-6 homologues, with the exception of the cod enzyme which was more active towards n-6 fatty acids.

We describe a polymorphism in the bovine gene PTHG which can be readily typed by PCR assay. The polymorphism is codominantly inherited and the allele frequencies appear characteristic of Bo indicus and B. taurus cattle.

Human Chromosome (Chr) 18 (HSA18) contains approximately85 million base pairs of DNA (Morton 1991). Thirty genes andpseudogenes have been mapped to HSA18 by somatic cell geneticsor in situ hybridization (Le Beau et al. 1993; Overhauser et al.1993; van Kessel et al. 1994). They include pituitary adenylcyclaseactivating peptide (ADCYAP1), laminin a chain (LAMA1), andmitochondrial NADH-ubiquinone reductase, 24 kDa subunit(NDUFV2) on the short arm, and transthyretin (TTR), B-cell lym-phoma 2 (BCL2), plasminogen activator inhibitor type 2 (PAI2),n-cadherin (CDH2), desmocollin type 2 (DSC2), ferrochelatase(FECH), cytochrome b5 (CYB5) and myelin basic protein (MBP)on the long arm (Fig. 1a). The murine homologs of HSA18 geneshave been mapped to at least four chromosomes as follows: Bcl-2and Planh2 genes are on MMU1, thymidylate synthase (Ts1) geneis on MMU5, Lama1 is on MMU17, and the rest are on MMU18(Mouse Genome Database 3.1, 1996).Two bovine genes—Yamaguchi sarcoma viral oncogene ho-molog 1 (YES1) and desmocollin type 1 (DSC1)—homologous toHSA18 loci have been mapped by in situ hybridization to Chr 24(Chowdhary et al. 1996), and ZOO-FISH analyses show thatHSA18 loci are located entirely on BTA24 (Hayes 1995; Toldo etal. 1995; Chowdhary et al. 1996). We report here the genomiclocalization of eight bovine genes homologous to HSA18 genesADCYAP1, CDH2, CYB5, DSC2, FECH, NDUFV2, PAI2, andTTR, using somatic cell genetics and linkage analysis in the In-ternational Bovine Reference Panel (IBRP).Oligonucleotide primers for PCR were designed on the basisof the published nucleotide sequences of sheep or cattle genesADCYAP1, CDH2, CYB5, DSC2, FECH, NDUFV2 and TTR, andwere used to amplify the corresponding DNA fragments frombovine total genomic DNA. Primers for PAI2 gene fragments weredesigned on the basis of a consensus sequence of the human andmurine genes. PCR amplification and single-strand conformationpolymorphism (SSCP) analysis were performed as described pre-viously (Agaba et al. 1996) except that the annealing and geltemperatures were optimized for each DNA fragment as shown inTable 1. The primers for PAI2 were allelic; that is, under optimumPCR conditions amplification of one allele from some individualswas favored over the alternative (null) allele, and, as expected,dominant inheritance was observed in informative IBRP families.Somatic cell hybrid mapping was achieved by PCR amplifica-tion of bovine sequences in a panel of 27 bovine × rodent somaticcell hybrids kindly provided by J. Womack of TexasA&MUniversity. In the case of the PAI2 gene, bovine fragments weredifferentiated from rodent fragments by SSCP analysis. Sevengenes, ADCYAP1, CDH2, CYB5, DSC2, FECH, PAI2 and TTR,segregated 100% concordantly with each other and with Chr 24,while the NDUFV2 gene segregated 100% concordantly with

Interspecies hybridization of bovids occurs between domestic cattle and at least 3 other species: the American bison (Bison bison), yak (Bos grunniens), and banteng (Bos banteng). Birth of a cattle � buffalo hybrid was reported in Russia, but the report was never authenticated. Such hybrids could be important in improving livestock production and managing diseases that impede production in tropical Africa. We investigated hybridization between cattle and their closest African wild relative, the African buffalo (Syncerus caffer caffer). In an attempt to produce pre-implantation cattle � buffalo hybrid embryos in vitro, matured bovine oocytes were subjected to a standard IVF procedure with either homologous (n = 1166 oocytes) or heterologous (n = 1202 oocytes) buffalo epididymal sperm. After IVF, 67.2% of the oocytes inseminated with homologous sperm cleaved. In contrast, insemination with buffalo sperm resulted in a 4.6% cleavage rate. Cleavage was also slower in hybrids than in cattle embryos. Up to 52.2% of cleaved homologous embryos progressed to the morula stage compared with 12.7% for hybrids. No hybrid embryos developed beyond the 16-cell stage, whereas 40.1% of the cleaved bovine embryos developed to the blastocyst stage. Developmental anomalies such as polyspermy, uneven cleavage, vacuolization, and absence of nuclei in some blastomeres were common in the hybrid embryos. We conclude that interspecies fertilization of cattle oocytes with African buffalo sperm occurs in vitro and that the barrier to hybridization is in the early stages of embryonic development. Also, chromosomal disparity is the likely cause of fertilization abnormalities, abnormal development, and subsequent arrest, impairing the formation of pre-implantation hybrid embryos. Investigation into developmental abnormalities, including reciprocal hybridization and genetic studies of the hybrid embryos, are recommended.

We have identified four single-strand conformation variants of the bovine tumor necrosis factor alpha gene by analysis of PCR-amplified fragments. The variants are inherited in Mendelian fashion and are informative for linkage mapping. We have mapped the bovine gene to Chromosome (Chr) 23 in a panel of somatic cell hybrids and observed genetic linkage to the major histocompatibility complex (BoLA) genes and microsatellite markers on bovine Chr 23 in an international bovine reference family panel. The distribution of the alleles was determined in cattle of different breeds and of different geographical origins, which included trypano-susceptible and trypano-tolerant cattle.

A cattle genetic linkage map was constructed which covers more than 95 percent of the bovine genome at medium density. Seven hundred and forty six DNA polymorphisms were genotyped in cattle families which comprise 347 individuals in full sibling pedigrees. Seven hundred and three of the loci are linked to at least one other locus. All linkage groups are assigned to chromosomes, and all are orientated with regards to the centromere. There is little overall difference in the lengths of the bull and cow linkage maps although there are individual differences between maps of chromosomes. One hundred and sixty polymorphisms are in or near genes, and the resultant genome-wide comparative analyses indicate that while there is greater conservation of synteny between cattle and humans compared with mice, the conservation of gene order between cattle and humans is much less than would be expected from the conservation of synteny. This map provides a basis for high-resolution mapping of the bovine genome with physical resources such as Yeast and Bacterial Artificial Chromosomes as well as providing the underpinning for the interpolation of information from the Human Genome Project.

The imprints of domestication and breed development on the genomes of livestock likely differ from those of companion animals. A deep draft sequence assembly of shotgun reads from a single Hereford female and comparative sequences sampled from six additional breeds were used to develop probes to interrogate 37,470 single-nucleotide polymorphisms (SNPs) in 497 cattle from 19 geographically and biologically diverse breeds. These data show that cattle have undergone a rapid recent decrease in effective population size from a very large ancestral population, possibly due to bottlenecks associated with domestication, selection, and breed formation. Domestication and artificial selection appear to have left detectable signatures of selection within the cattle genome, yet the current levels of diversity within breeds are at least as great as exists within humans.