Seven novel KIT mutations in horses with white coat colour phenotypes

White coat colour in horses is inherited as a monogenic autosomal dominant trait showing a variable expression of coat depigmentation. Mutations in the KIT gene have previously been shown to cause white coat colour phenotypes in pigs, mice and humans. We recently also demonstrated that four independent mutations in the equine KIT gene are responsible for the dominant white coat colour phenotype in various horse breeds. We have now analysed additional horse families segregating for white coat colour phenotypes and report seven new KIT mutations in independent Thoroughbred, Icelandic Horse, German Holstein, Quarter Horse and South German Draft Horse families. In four of the seven families, only one single white horse, presumably representing the founder for each of the four respective mutations, was available for genotyping. The newly reported mutations comprise two frameshift mutations (c.1126_1129delGAAC; c.2193delG), two missense mutations (c.856G>A; c.1789G>A) and three splice site mutations (c.338-1G>C; c.2222-1G>A; c.2684+1G>A). White phenotypes in horses show a remarkable allelic heterogeneity. In fact, a higher number of alleles are molecularly characterized at the equine KIT gene than for any other known gene in livestock species.     

Effect of Iron on Erythropoietin Production in Anaemic Piglets

In the present study the response of plasma erythropoietin to iron injection in anaemic piglets was examined. At the age of 16 days, 4 piglets from the same litter were given 180 mg iron subcutane-ously. After iron injection, blood samples for estimation of erythropoietin activity in plasma, haemoglobin concentration, and reticulocyte counts were taken every 6 or 12 h for 3 Vi days. Plasma erythropoietin activity was estimated by a monoclonal enzyme-im-munoassay (ELISA), developed for human erythropoietin. On the day of iron injection, haemoglobin concentration ranged between 41 and 48 g/1, reticulocyte counts from 9 to 17 percent, and plasma erythropoietin between 22 and 144 mU/ml. In 3 of the 4 piglets, iron injection resulted in a 2-6 fold increase in erythropoietin activity. Maximal eryth-ropoietin activities were observed 24-42 h after injection, and after 66 h, the activities were close to the pretreatment values. It is concluded that in our experiment, iron, per se, has stimulated erythropoietin production.     

白色基因座(I)在中国地方猪种毛色遗传中的作用研究

通过利用PCR—RFLP和PCR—SSCP技术对中国地方猪种KIT基因内含子17、18的序列进行多态性分析。结果表明：内含子17上的替换突变(G→A)发生于毛色为白色的个体——白色五指山猪、大白猪、长白猪上,其基因型(AB型)频率分别为1、1和0．8;其他中国地方猪种的此基因型频率均为0。内含子18上的缺失突变(AGTT)也同样发生在上述3个猪种的白色个体中,其基因型(AA型)频率分别为1、1和0．93;而且同样在其他的地方品种中其基因型频率均为0。这充分证明KIT基因对于猪的白毛色有重要的调控作用,而且I基因座对于其他的经典遗传基因座有上位作用。另一方面,中国地方猪种荣昌猪虽然在表型上与引入猪种大白猪、长白猪相似(白毛色),但是在KIT基因上发生的突变完全不同,推测它们分别属于不同的毛色遗传体系。     

Molecular genetics of coat colour variations in White Galloway and White Park cattle

White Galloway cattle exhibit three different white coat colour phenotypes, that is, well marked, strongly marked and mismarked. However, mating of individuals with the preferred well or strongly marked phenotype also results in offspring with the undesired mismarked and/or even fully black coat colour. To elucidate the genetic background of the coat colour variations in White Galloway cattle, we analysed four coat colour relevant genes: mast/stem cell growth factor receptor (KIT), KIT ligand (KITLG), melanocortin 1 receptor (MC1R) and tyrosinase (TYR). Here, we show that the coat colour variations in White Galloway cattle and White Park cattle are caused by a KIT gene (chromosome 6) duplication and aberrant insertion on chromosome 29 (Cs₂₉) as recently described for colour‐sided Belgian Blue. Homozygous (Cs₂₉/Cs₂₉) White Galloway cattle and White Park cattle exhibit the mismarked phenotype, whereas heterozygous (Cs₂₉/wt₂₉) individuals are either well or strongly marked. In contrast, fully black individuals are characterised by the wild‐type chromosome 29. As known for other cattle breeds, mutations in the MC1R gene determine the red colouring. Our data suggest that the white coat colour variations in White Galloway cattle and White Park cattle are caused by a dose‐dependent effect based on the ploidy of aberrant insertions and inheritance of the KIT gene on chromosome 29.     

Linked markers exclude KIT as the gene responsible for appaloosa coat colour spotting patterns in horses

The appaloosa coat colour pattern of the horse is similar to that caused by the rump-white (Rw) gene in the mouse. In the mouse Rw colour pattern is the result of an inversion involving the proto-oncogene c-kit (KIT). Therefore, we investigated KIT as a candidate gene that encodes the appaloosa coat colour gene (Lp) in horses. KIT plays a critical role in haematopoiesis, gametogenesis, and melanogenesis and encodes a transmembrane tyrosine kinase receptor that belongs to the PDGF/CSF-1/c-KIT receptor subfamily. Half-sib families segregating for Lp were uninformative for a reported polymorphism in KIT. However, KIT is located on horse chromosome 3 close to albumin (ALB), serum carboxylesterase (ES), vitamin D-binding protein (GC) and microsatellite markers ASB23, LEX007, LEX57, and UCDEQ437. Indeed, KIT and ASB23 were localized to ECA3q21-22.1 and 3q22.1-22.3, respectively, by fluorescent in situ hybridization. Family studies were conducted to investigate linkage of Lp to these markers using eight half-sib families in which Appaloosa stallions were mated to solid coloured mares. Linkage of Lp to the chromosome region containing ES, ALB, GC, ASB23, UCDEQ437, LEX57, and LEX007 was investigated by a multipoint linkage analysis using the computer program GENEHUNTER. LOD scores over the interval under investigation ranged from -4.28 to -12.48, with a score of -12.48 at the location for ASB23. Therefore, it was concluded that appaloosa (Lp) is not linked to any of the tested markers on ECA3, and thus Lp is unlikely to be the product of KIT.     

The Ratio of sTfR/Ferritin is Associated with the Expression Level of TfR in Rat Bone Marrow Cells After Endurance Exercise

Currently, it is unclear which index of haematological parameters could be used to most easily monitor iron deficiency during endurance training. To address this question, 16 male Wistar rats were randomly assigned to two groups: a sedentary group (n = 8) and an exercised group (n = 8). Initially, animals in the exercise group started running on a treadmill at a rate of 30 m/min, on a 0% grade, for 1 min/session. Running time was gradually increased by 2 min/day. The training plan was one session per day during the initial 2 weeks and two sessions per day during the third to ninth week. At the end of the 9-week experiment, we analysed the blood of the experimental animals for haemoglobin levels, erythrocyte numbers, haematocrit, serum iron levels, total iron binding capacity, transferrin saturation, serum ferritin levels and soluble transferrin receptor (sTfR) levels, and we calculated the ratio of sTfR/ferritin. Erythrocyte numbers, haemoglobin levels and haematocrit values were decreased after 9 weeks of exercise, but sTfR and sTfR/ferritin values were increased (P < 0.01 or P < 0.05). The training regime significantly increased TfR mRNA levels in the bone marrow cells of the exercised rats compared with the sedentary group (1.8 ± 0.5 vs. 1.1 ± 0.2, P < 0.01). These results revealed a significant correlation between TfR levels in the bone marrow cells and the ratio of sTfR/ferritin (r = 0.517; P < 0.01) and sTfR levels (r = 0.206; P < 0.05) in sedentary and exercised rats. In conclusion, we show that sTfR indices and the ratio of sTfR/ferritin could be useful indicators for monitoring iron deficiency during endurance training.     

Association of BF,RBP4, and ESR2 Genotypes with Litter Size in an Autochthonous Pig Population

The aim of the present study was to determine any potential association of the BF, RBP4, and ESR2 genes with reproduction traits in an autochthonous Greek pig population. The PCR-RFLP methodology was implemented for genotyping purposes of the examined genes. No deviation from the Hardy-Weinberg equilibrium was observed for the examined loci, while the B allele noted to be the more frequent in all analyzed genes. In addition, sows with the AA genotype of BF gene found to produce significantly lower numbers of the total born piglets (TNB) and number of piglets born alive (TNA), while the respective BB genotype significantly exceeded in TNB and NBA traits compared to the other two genotypes (P?Abbreviations: TNB: Total number of born piglets; NBA: Number of piglets born alive     

Oxygen transport to tissue modified by entrapment of an allosteric effector of haemoglobin in erythrocytes

Oxygen affinity of haemoglobin is modulated by several parameters such as the allosteric effector 2-3 DPG for most mammalians. Inositol hexaphosphate (I.H.P.) exerts the same effect on haemoglobin. A previously developed new methodology for the entrapment of drugs into erythrocytes has been adapted to I.H.P.; it is based on a reversible osmotic shock. I.H.P. loaded red blood cells have characteristics very similar to those of native cells. The decrease in oxygen affinity is related to the dose of encapsulated I.H.P. In piglets, transfusion of such cells has led to an increase of oxygen extraction from haemoglobin. Normal anesthetized animals regulate their oxygen consumption by reduction of cardiac output.     

A large duplication associated with dominant white color in pigs originated by homologous recombination between LINE elements flanking KIT

The Dominant White (I/KIT) locus is one of the major coat color loci in the pig. Previous studies showed that the Dominant White (I) and Patch (IP) alleles are both associated with a duplication including the entire KIT coding sequence. We have now constructed a BAC contig spanning the three closely linked tyrosine kinase receptor genes PDGFRA–KIT–KDR. The size of the duplication was estimated at about 450 kb and includes KIT, but not PDGFRA and KDR. Sequence analysis revealed that the duplication arose by unequal homologous recombination between two LINE elements flanking KIT. The same unique duplication breakpoint was identified in animals carrying the I and IP alleles across breeds, implying that Dominant White and Patch alleles are descendants of a single duplication event. An unexpected finding was that Piétrain pigs carry the KIT duplication, since this breed was previously assumed to be wild type at this locus. Comparative sequence analysis indicated that the distinct phenotypic effect of the duplication occurs because the duplicated copy lacks some regulatory elements located more than 150 kb upstream of KIT exon 1 and necessary for normal KIT expression.     

Confirmation of QTL on porcine chromosomes 1 and 8 influencing leukocyte numbers, haematological parameters and leukocyte function

A genome wide search in European Wild Boar x Swedish Yorkshire (W x Y) inter-cross pigs has earlier identified quantitative trait loci (QTL) for leucocyte number and function on porcine chromosomes 1 and 8 (SSC 1 and 8). To verify the involvement of these chromosomal regions in the regulation of haematocrit (Hem) and haemoglobin (Hb) levels, leucocyte numbers and in vitro leukocyte functions (mitogen induced proliferation and IL-2 production, virus induced interferon-alpha production and neutrophil phagocytosis), animals of different genetic backgrounds were analysed. The animals comprised a back-cross sire family (n=47) of W x Y pigs and six crossbred [Y x Landrace (L)] sire families (n=191). They were genotyped for 16 genetic markers and an interval analysis was performed. On SSC1, a QTL close to S0082 on the q-arm that influenced numbers of white blood cells in L x Y pigs and numbers of band neutrophils and CD8(+) cells in W x Y pigs was identified (P相似文献   

Dietary ractopamine supplementation of pregnant sows: what are the impacts on the neonate?

The use of additives such as ractopamine (Rac) in pregnant sows during early-mid pregnancy is an alternative to increase foetal and progeny growth and development. However, Rac supplementation in finishing pigs can lead to behavioural and physiological changes similar to the typical stress responses. The objective of this study was to evaluate the effects of dietary supplementation with Rac in pregnant sows from day 25 to 50 of gestation (pre-hyperplastic stage) on piglet’s vitality, blood parameters, number, diameter and perimeter of muscle fibres in semitendinosus muscle and developmental characteristics of piglets at birth to weaning. Forty-one hybrid sows were divided into three dietary treatments: (1) control diet without Rac (control), (2) addition of 10 mg/kg of Rac (Rac10) and (3) addition of 20 mg/kg of Rac (Rac20). Higher numbers of low-vitality piglets (P<0.05) were observed in Rac-fed sows, regardless of dose, compared with the control group. Very low-density lipoprotein levels were lower in the Rac10 group when compared with the Rac20 group at day 21. Haematocrit was greater, and the mean corpuscular haemoglobin concentration was lower in piglets from Rac-fed sows. No significant statistical differences were detected regarding piglets body weight, average daily gain, blood gasometry, complete blood count and muscle fibre measurements in semitendinosus muscle. The use of Rac in pregnant sows reduced the vitality parameters of piglets but did not improve the performance from birth until weaning and did not negatively influence the haematological parameter and lipid metabolism.     

Relationship of season, thermal loading and red-sore disease with various haematological parameters in Micropterus salmoides

One hundred and fifty largemouth bass (Micropterus salmoides) were caught during the mid-winter and mid-summer months in Par Pond, an 1120 ha cooling reservoir located on Savannah River Plant near Aiken, SC. Equal numbers of fish were taken from ambient and thermally altered (> 10° C ambient) areas of the lake. The total length and weight of each bass was noted, along with the presence or absence of lesions associated with red-sore disease (caused by the bacterium, Aeromonas hydrophila and the ciliate, Epistylis sp.). Blood was drawn, by heart puncture, from each bass and the following haematological parameters measured: haematocrit, haemoglobin, iron, total iron-binding capacity, thyroxine, sodium and potassium. Only the total iron-binding capacity was significantly higher in bass with red-sore disease; none of the other blood parameters were significantly different in infected and uninfected bass. Bass from thermally altered locations had consistently higher haematocrits during both summer and winter. Bass from thermal and ambient locations were not different with respect to iron, iron-binding capacity, sodium or potassium levels. However, bass taken during summer months had higher levels of circulating iron than individuals in winter months. Summer bass had higher thyroxine levels than bass in winter. These results are discussed in terms of their relationship to red-sore disease, season and the impact of thermal effluent.     

Chinese white Rongchang pig does not have the dominant white allele of KIT but has the dominant black allele of MC1R

The mast/stem cell growth factor receptor (KIT) and melanocortin receptor 1 (MC1R) mutations are responsible for coat color phenotypes in domestic pigs. Rongchang is a Chinese indigenous pig breed with a white coat color phenotype. To investigate the genetic variability of the KIT and MC1R genes and their possible association with the coat color phenotype in this breed, a gene duplication and splice mutation of KIT were diagnosed in a sample of 93 unrelated Rongchang animals. The results show that Rongchang pigs have a single copy of KIT without the splice mutation at the first nucleotide of intron 17, indicating that the dominant white I allele of KIT is not responsible for their white phenotype. The KIT mRNA and MC1R coding sequences were also determined in this breed. Three putative amino acid substitutions were found in the KIT gene between Rongchang and Western white pigs, their association with the Rongchang white phenotype remains unknown. For the MC1R gene, Rongchang pigs were demonstrated to have the same dominant black allele (E(D1)) as other Chinese breeds, supporting the previous conclusion that Chinese and Western pigs have independent domestication origin. We also clarified that the Rongchang white phenotype was recessive to nonwhite color phenotypes. Our results provide a good starting point for the identification of the mutations underlying the white coat color in Rongchang pigs.     

Comparative study of some haematological and biochemical blood parameters in fishes from the Skagerrak

Marine fishes caught in the Skagerrak, 27 different species representing various groups of fishes (Cyclostomi, Holocephali, Elasmobranchii and Teleostei), were examined for the following haematological and biochemical blood parameters: haematocrit, haemoglobin, mean corpuscle haemoglobin concentration, total plasma protein, blood glucose and blood lactate. Interspecies variations as well as variations within some species were observed. The haemoglobin values for all species showed a positive correlation to the corresponding haematocrit values. Relatively low values for haematocrit and haemoglobin were found in cyclostomes, holocephaleans and elasmobranchii compared to the majority of teleosts. Within the teleost group, the haematocrit and haemoglobin levels were positively correlated with the activity of the fish species. The cyclostome Myxine glutimsa L. had a total plasma protein content in the same range as most teleosts, whereas holocephaleans, elasmobranchii and the deep-water teleost Coryphaenoides rupestris Gunnerus showed comparatively low values. Among teleosts some relationship seemed to exist between the total plasma protein level and the activity of the fish species. In addition, a correlation between plasma protein content and levels of blood lipids were noted. Values for blood glucose and blood lactate were found to be lower in cyclostomes, holocephaleans and elasmobranchii than in most teleosts. Higher blood glucose levels were observed in the more active teleost species.     

Anaemia in Housed Lambs: Effects of Oral Iron on Clinical Pathology and Performance

An experiment including 39 pairs of housed twin lambs was performed to evaluate the effect of an oral iron supplement (Fe-MAX® Starter) on clinical pathology, growth rates and disease occurrence. Significant differences between the iron supplemented group (Fe-group) and the controls were seen, for varying periods of time, for all red blood cell and iron parameters examined. In spite of this, 25% of the iron supplemented lambs had haemoglobin values below 80 g/L 4 weeks after treatment, whereas 33% of the controls had corresponding values 3 weeks after treatment, indicating that one single iron dose was insufficient to prevent iron deficiency anaemia. No significant positive effect on live weights was seen. However, the Fe-group had a poorer daily weight gain during the first week after dosing (p<0.01), but a better daily gain during the second (p<0.01) and third weeks. No effects were seen on disease occurrence. An additional trial with iron dextran injections to 5 untreated 21 days old lambs with varying haemoglobin values, revealed a rapid increase in the red cell distribution in anaemic lambs, with production of macrocytic, mainly normochromic erythrocytes. The new erythrocyte population was visible on the cytogram after 2 to 3 days and on the histogram after 5 days.     

Enterococcus faecium EK13--an enterocin a-producing strain with probiotic character and its effect in piglets

The experiment was conducted to determine the effects of the inoculation of the probiotic and enterocin A-producing strain Enterococcus faecium EK13 on selected parameters of metabolic profile, gut microflora, growth, and health in newborn piglets of Slovak White Improved. Piglets for study were divided into two groups: one group (EK13 group, n=8) received strain EK13 per os once daily for 7 days (2ml per piglet, 10(9)CFU/mL of saline buffer). The control group of piglets (n=7) was given placebo-saline buffer. The experiment lasted 14 days. After 7 days, strain EK13 reached 9.8 log(10) CFU/g in faeces of E. faecium EK13 treated piglets while counts of Escherichia coli were significantly lower (P<0.01) than in piglets of the control group. The concentrations of total serum protein, calcium, haemoglobin, haematocrit, red blood cell count and index of phagocytic activity of leukocytes were significantly higher after application of strain EK13. On the other hand, cholesterol was significantly lower in the EK13 group of animals. On day 14, piglets were killed and samples of intestinal contents were taken. Total counts of bacteria in the intestinal contents (jejunum, ileum, caecum, colon) were not significantly influenced. The pH value was significantly lower (P<0.05) only in duodenum of piglets receiving E. faecium EK13. There was a significant higher concentration of lactic acid (P<0.01) and propionic acid in the colon (P<0.001) of the EK13 group. Application of E. faecium EK13 did not influence the daily body weight gain significantly.     

A defective seed coat pattern (Net) is correlated with the post-transcriptional abundance of soluble proline-rich cell wall proteins

The pigmented seed coats of several soybean (Glycine max (L.) Merr.) plant introductions and isolines have unusual defects that result in cracking of the mature seed coat exposing the endosperm and cotyledons. It has previously been shown that the T (tawny) locus that controls the color of trichomes on stems and leaves also has an effect on both the structure and pigmentation of the seed coat. Distribution of pigmentation on the seed coat is controlled by alleles of the I (inhibitor) locus. It was also found that total seed coat proteins were difficult to extract from pigmented seed coats with i T genotypes because they have procyanidins that exhibit tannin properties. We report that the inclusion of poly-L-proline in the extraction buffer out-competes proteins for binding to procyanidins. Once this problem was solved, we examined expression of the proline-rich cell wall proteins PRP1 and PRP2 in pigmented genotypes with the dominant T allele. We found that both homozygous i T and i t genotypes have reduced soluble PRP1 levels. The epistatic interaction of the double recessive genotype at both loci is necessary to produce the pigmented, defective seed coat phenotype characteristic of seed coats with the double recessive i and t alleles. This implies a novel effect of an enzyme in the flavonoid pathway on seed coat structure in addition to its effect on flavonoids, anthocyanidins, and proanthocyanidins. No soluble PRP1 polypeptides were detectable in pigmented seed coats (i T genotypes) of isolines that also display a net-like pattern of seed coat cracking, known as the Net defect. PRP2 was also absent in one of the these lines. However, both PRP1 and PRP2 cytoplasmic mRNAs were found in the Net-defective seed coats. Together with in vitro translation studies, these results suggest that the absence of soluble PRP polypeptides in the defective Net lines is post-translational and could be due to a more rapid or premature insolubilization of PRP polypeptides within the cell wall matrix.     

Methanol-fermentation protein in veal calf nutrition

In an experiment with 4 × 12 male veal calves, the influence of 0, 2.5, 5.0 and 7.5% ICI protein and sweet whey powder in a milk replacer on growth; feed efficiency; blood concentrations of urea, uric acid and allantoin; and on the iron status, colour of the meat and carcass grade of the animals was investigated. The proportion of skimmed milk powder in the milk replacer was thus reduced in protein equivalence by 5, 10 and 15% (groups II–IV).Feeding with 2.5, 5 and 7.5% ICI protein in the milk replacer gave, over the whole fattening period (60–150 kg), about the same rates of growth and feed efficiency as the exclusive use of milk protein. While the urea content in the serum was not significantly influenced by the protein sources used, the uric acid and allantoin levels of the groups given ICI protein had risen greatly at 5 weeks and at the end of fattening experiment. Despite the higher total iron supply to groups II, III and IV, the criteria indicating iron supply such as haemoglobin, haematocrit and myoglobin levels did not show any increase compared to group I. Similarly, no significant differences in meat colour and carcass grade were found among the experimental groups.For veal calves the level of ICI protein in the milk replacer can be up to 7.5% during the entire feeding period.     

Impact of the probiotic bacteria Enterococcus faecium NCIMB 10415 (SF68) and Bacillus cereus var. toyoi NCIMB 40112 on the development of serum IgG and faecal IgA of sows and their piglets

To examine the influence of two different probiotic bacteria on the humoral immune system of swine, two animal studies were carried out with sows and their litters. The sows' feed was supplemented with either Enterococcusfaecium NCIMB 10415 (SF68) or Bacillus cereus var. toyoi NCIMB 40112 beginning early in pregnancy. The total IgA content in the faeces as well as the total IgG concentration in the blood of the sows was recorded before and after weaning. The same parameters were determined in the blood and faeces of the piglets. In sows, only feed supplementation with B. cereus led to a clear increase in faecal IgA. Serum IgG levels were not significantly affected by any probiotic feeding in sows. In piglets, the group that was fed B. cereus showed significantly higher faecal IgA levels shortly before weaning, whereas in the E. faecium group, a significant decrease in IgA levels was observed one week after weaning. In both probiotic fed groups the post-weaning IgG levels were significantly decreased compared to the respective control groups. We conclude that B. cereus var. toyoi feed supplementation led to an increased intestinal IgA secretion both in sows and piglets. This effect could be related to a more successful mucosal defence which in turn led to a lower level in systemic IgG production in piglets after weaning.