Molecular characterization of the llamas (Lama glama) casein cluster genes transcripts (CSN1S1, CSN2, CSN1S2, CSN3) and regulatory regions

In the present paper, we report for the first time the characterization of llama (Lama glama) caseins at transcriptomic and genetic level. A total of 288 casein clones transcripts were analysed from two lactating llamas. The most represented mRNA populations were those correctly assembled (85.07%) and they encoded for mature proteins of 215, 217, 187 and 162 amino acids respectively for the CSN1S1, CSN2, CSN1S2 and CSN3 genes. The exonic subdivision evidenced a structure made of 21, 9, 17 and 6 exons for the αs1-, β-, αs2- and κ-casein genes respectively. Exon skipping and duplication events were evidenced. Two variants A and B were identified in the αs1-casein gene as result of the alternative out-splicing of the exon 18. An additional exon coding for a novel esapeptide was found to be cryptic in the κ-casein gene, whereas one extra exon was found in the αs2-casein gene by the comparison with the Camelus dromedaries sequence. A total of 28 putative phosphorylated motifs highlighted a complex heterogeneity and a potential variable degree of post-translational modifications. Ninety-six polymorphic sites were found through the comparison of the lama casein cDNAs with the homologous camel sequences, whereas the first description and characterization of the 5'- and 3'-regulatory regions allowed to identify the main putative consensus sequences involved in the casein genes expression, thus opening the way to new investigations -so far- never achieved in this species

Association study in naturally infected helminth layers shows evidence for influence of interferon-gamma gene variants on Ascaridia galli worm burden

Single nucleotide polymorphisms (SNPs) in the genes for interleukin-4, -13 and interferon-gamma, and 21 additional SNPs which previously had been significantly associated with immune traits in the chicken, were genotyped in white and brown layer hens and analyzed for their association with helminth burden following natural infections. A nucleotide substitution located upstream of the promoter of the interferon-gamma gene was significantly associated with the log transformed number of Ascaridia galli in the brown layer line (genotype CC: 6.4 ± 1.0 worms; genotype CT: 11.7 ± 2.2 worms). Therefore, IFNG seems to be a promising candidate gene for further studies on helminth resistance in the chicken

Use of molecular markers for evaluation of genetic diversity and in animal production

Genetic markers provide information about allelic variation at a given locus. The increasing availability of molecular markers in farm animals allows the detailed analyses and evaluation of genetic diversity and furthermore the detection of genes influencing economically important traits. The majority of molecular markers used nowadays with high-throughput systems are microsatellite markers (simple tandem repeat, STR). A variable number of tandem repeats (VNTRs), random amplified polymorphic DNA (RAPD) markers, single-strand conformation polymorphisms (SSCPs), restriction fragment length polymorphisms (RFLPs) and amplified fragment length polymorphisms (AFLPs) markers are not commonly applied in farm animals. Additionally within diversity and phylogeny studies specific mtDNA and Y chromosome markers are used for the identification of maternal and paternal lineages. Beyond diversity studies, molecular markers are also used for mapping quantitative trait loci (QTL) and within the marker-assisted selection (MAS). Until recently microsatellites were the markers used for mapping quantitative trait loci for production and functional traits in farm animals and tightly linked markers are used for marker-assisted selection in practice. They are also the prerequisite for the identification of positional and functional candidate genes responsible for quantitative traits. The detailed use of molecular markers for the evaluation of genetic diversity and identification of economically important traits in animal production is presented on the basis of different examples

High Polymorphism in the K-Casein (CSN3) Gene from Wild and Domestic Caprine Species Revealed by DNA Sequencing

We assessed polymorphisms in exon IV of the k-casein gene (CSN3) in ten different breeds of domestic goat (Capra hircus) from three continents and in three related wild caprine taxa (Capra ibex, Capra sibirica and Capra aegagrus). Thirty-five DNA samples were sequenced within a 558 bp fragment of exon IV. Nine polymorphic sites were identified in domestic goat, including four new polymorphisms. In addition to four previously described polymorphic positions, a total of 13 polymorphisms allowed the identification of 13 DNA variants, corresponding to 10 protein variants. Because of conflicting nomenclature of these variants, we propose a standardized allele designation. CSN3*A, CSN3*B, and CSN3*D were found as widely distributed alleles in European goat breeds. Within Capra ibex we identified three variants and showed that the sequence of Capra aegagrus is identical to the most common Capra hircus variant, consistent with Capra aegagrus being the wild progenitor of domestic goats. A dendrogram was drawn to represent the molecular network between the caprine CSN3 variants

Rapid testing leads to the underestimation of the scrapie prevalence in an affected sheep and goat flock

To obtain a more detailed understanding of the prevalence of classical scrapie infections in a heavily affected German sheep flock (composed of 603 sheep and 6 goats), we analysed 169 sheep and 6 goats that carried the genotypes susceptible to the disease and that were therefore culled following discovery of the index case. The initial tests were performed using the Biorad TeSeE ELISA and reactive results were verified by official confirmatory methods (OIE-immunoblot and/or immunohistochemistry (IHC)) to demonstrate the deposition of scrapie-associated PrPSc in the brain stem (obex). This approach led to the discovery of 40 additional subclinically scrapie-infected sheep. Furthermore, peripheral lymphatic and nervous tissue samples of the 129 sheep and 6 goats with a negative CNS result were examined by IHC in order to identify any preclinical infections which had not already spread to the central nervous system (CNS). Using this approach we found 13 additional sheep with PrPSc depositions in the gut-associated lymph nodes (GALT) as well as in the enteric nervous system. Moreover, in most of these cases PrPSc was also deposited in the spleen and in the retropharyngeal and superficial cervical lymph nodes. Taken together, these results show a 30.3% infection prevalence in this scrapie-affected flock. Almost 7.4% of the infected animals harboured PrPSc exclusively in the peripheral lymphatic and nervous tissue and were therefore missed by the currently used testing strategy

Genes involved in carnitine synthesis and carnitine uptake are up-regulated in the liver of sows during lactation

BACKGROUND:Convincing evidence exist that carnitine synthesis and uptake of carnitine into cells is regulated by peroxisome proliferator-activated receptor alpha (PPARA), a transcription factor which is physiologically activated during fasting or energy deprivation. Sows are typically in a negative energy balance during peak lactation. We investigated the hypothesis that genes involved in carnitine synthesis and uptake in the liver of sows are up-regulated during peak lactation. FINDINGS:Transcript levels of several PPARalpha target genes involved in fatty acid uptake (FABP4, SLC25A20), fatty acid oxidation (ACOX1, CYP4A24) and ketogenesis (HMGCS2, FGF21) were elevated in the liver of lactating compared to non-lactating sows (P < 0.05). In addition, transcript levels of genes involved in carnitine synthesis (ALDH9A1, TMLHE, BBOX1) and carnitine uptake (SLC22A5) in the liver were greater in lactating than in non-lactating sows (P < 0.05). Carnitine concentrations in liver and plasma were about 20% and 50%, respectively, lower in lactating than in non-lactating sows (P < 0.05), which is likely due to an increased loss of carnitine via the milk. CONCLUSIONS:The results of the present study show that PPARalpha is activated in the liver of sows during lactation which leads to an up-regulation of genes involved in carnitine synthesis and carnitine uptake. The PPARalpha mediated up-regulation of genes involved in carnitine synthesis and uptake in the liver of lactating sows may be regarded as an adaptive mechanism to maintain hepatic carnitine levels at a level sufficient to transport excessive amounts of fatty acids into the mitochondrion

Treatment of lactating sows with clofibrate as a synthetic agonist of PPARalpha does not influence milk fat content and gains of litters

BACKGROUND: In rats, it has been observed that treatment with activators of peroxisome proliferator-activated receptor a (PPARalpha) disturbs metabolic adaptations during lactation, which in turn lead to a reduction of milk fat content and gains of litters during the suckling period. It has not yet been investigated whether agonists of PPARalpha are impairing milk production of lactating sows in a similar manner as in rats. Therefore, the present study aimed to investigate the effect of treatment with clofibrate, a strong synthetic agonist of PPARalpha, on milk composition and litter gains in lactating sows. RESULTS: Twenty lactating sows received either a basal diet (control group) or the same diet with supplementation of 2 g of clofibrate per kg of diet (clofibrate group). In the clofibrate group, mRNA concentrations of various PPARalpha target genes involved in fatty acid utilization in liver and skeletal muscle were moderately up-regulated. Fat and energy content of the milk and gains of litters during the suckling period were not different between the control group and the clofibrate group. CONCLUSIONS: It is shown that treatment with clofibrate induces only a moderate up-regulation of PPARalpha target genes in liver and muscle of lactating sows and in turn might have limited effect on whole body fatty acid utilization. This may be the reason why clofibrate treatment did not influence milk fat content and gains of litters during the suckling period. Thus, the present study indicates that activation of PPARalpha induced either by native agonists such as dietary polyunsaturated fatty acids or a by negative energy balance might be largely uncritical in lactating sows with respect to milk production and litter gains in lactating sows

Geographic distribution of haplotype diversity at the bovine casein locus

The genetic diversity of the casein locus in cattle was studied on the basis of haplotype analysis. Consideration of recently described genetic variants of the casein genes which to date have not been the subject of diversity studies, allowed the identification of new haplotypes. Genotyping of 30 cattle breeds from four continents revealed a geographically associated distribution of haplotypes, mainly defined by frequencies of alleles at CSN1S1 and CSN3. The genetic diversity within taurine breeds in Europe was found to decrease significantly from the south to the north and from the east to the west. Such geographic patterns of cattle genetic variation at the casein locus may be a result of the domestication process of modern cattle as well as geographically differentiated natural or artificial selection. The comparison of African Bos taurus and Bos indicus breeds allowed the identification of several Bos indicus specific haplotypes (CSN1S1*C-CSN2*A2-CSN3*AI/CSN3*H) that are not found in pure taurine breeds. The occurrence of such haplotypes in southern European breeds also suggests that an introgression of indicine genes into taurine breeds could have contributed to the distribution of the genetic variation observed

Genes involved in carnitine synthesis and carnitine uptake are up-regulated in the liver of sows during lactation

Abstract Background: Convincing evidence exist that carnitine synthesis and uptake of carnitine into cells is regulated by peroxisome proliferator-activated receptor α (PPARA), a transcription factor which is physiologically activated during fasting or energy deprivation. Sows are typically in a negative energy balance during peak lactation. We investigated the hypothesis that genes involved in carnitine synthesis and uptake in the liver of sows are upregulated during peak lactation. Findings: Transcript levels of several PPARα target genes involved in fatty acid uptake (FABP4, SLC25A20), fatty acid oxidation (ACOX1, CYP4A24) and ketogenesis (HMGCS2, FGF21) were elevated in the liver of lactating compared to non-lactating sows (P &lt; 0.05). In addition, transcript levels of genes involved in carnitine synthesis (ALDH9A1, TMLHE, BBOX1) and carnitine uptake (SLC22A5) in the liver were greater in lactating than in non-lactating sows (P &lt; 0.05). Carnitine concentrations in liver and plasma were about 20% and 50%, respectively, lower in lactating than in non-lactating sows (P &lt; 0.05), which is likely due to an increased loss of carnitine via the milk. Conclusions: The results of the present study show that PPARα is activated in the liver of sows during lactation which leads to an up-regulation of genes involved in carnitine synthesis and carnitine uptake. The PPARα mediated up-regulation of genes involved in carnitine synthesis and uptake in the liver of lactating sows may be regarded as an adaptive mechanism to maintain hepatic carnitine levels at a level sufficient to transport excessive amounts of fatty acids into the mitochondrion