Differential cytokine expression in T cell subsets from bovine peripheral blood

Although distinct cytokine expression in T cell subsets is well understood in mice and humans, limited information is available on bovine T cell subsets. In the present study, we analyzed the mRNA expression of 10 kinds of cytokines and CD25 expression in CD4⁺, CD8⁺, WC1⁺ and WC1^-γδ T cell subsets in bovine peripheral blood by Concanavalin A (Con A) stimulation. CD25 expression was significantly increased in CD4⁺, CD8⁺ and WC1⁺γδ T cells, but not in WC1^-γδ T cells by Con A stimulation. In CD4⁺ T cells, the mRNAs of Interleukin (IL)-2, IL-6, IL-10, interferon (IFN)-γ, tumor necrosis factor (TNF)-α, TNF-β and transforming growth factor (TGF)-β were expressed in control cultures, and IL-3, IL-4 and granulocyte-macrophage colony stimulating factor (GM-CSF) were newly expressed when the cells were stimulated with Con A. CD8⁺ T cells expressed the mRNAs of IL-6, TNF-α, TNF-β and TGF-β in control cultures, and newly expressed those of IL-2, IFN-γ and GM-CSF, but did not express those of IL-3, IL-4 or IL-10 after Con A stimulation. The cytokine expression profile of WC1⁺γδ T cells was similar to that of CD8⁺ T cells. However, WC1^-γδ T cells did not express any cytokine mRNA except TGF-â mRNA. These results will contribute to elucidate the participation of T cell subsets in immune responses against infectious disease in cattle.     

IARS mutation causes prenatal death in Japanese Black cattle

Isoleucyl‐tRNA synthetase (IARS) c.235G > C (p.V79L) is a causative mutation for a recessive disease called IARS disorder in Japanese black cattle. The disease is involved in weak calf syndrome and is characterized by low birth weight, weakness and poor suckling. The gestation period is often slightly extended, implying that intrauterine growth is retarded. In a previous analysis of 2597 artificial insemination (AI) procedures, we suggested that the IARS mutation might contribute toward an increase in the incidence of prenatal death. In this study, we extended this analysis to better clarify the association between the IARS mutation and prenatal death. The IARS genotypes of 92 animals resulting from crosses between carrier (G/C) × G/C were 27 normal (G/G), 55 G/C and 10 affected animals (C/C) (expected numbers: 23, 46 and 23, respectively). Compared to the expected numbers, there were significantly fewer affected animals in this population (P < 0.05), suggesting that more than half of the affected embryos died prenatally. When the number of AI procedures examined was increased to 11 580, the frequency of re‐insemination after G/C × G/C insemination was significantly higher at 61‐140 days (P < 0.001). The findings suggested that the homozygous IARS mutation not only causes calf death, but also embryonic or fetal death.     

Identification of the core rumen bacterial taxa and their population dynamics during the fattening period in Japanese Black cattle

The rumen microbiota comprises a vast range of bacterial taxa, which may affect the production of high-quality meat in Japanese Black cattle. The aim of this study was to identify core rumen microbiota in rumen fluid samples collected from 74 Japanese Black cattle raised under different dietary conditions using 16S rRNA gene amplicon sequencing. In the rumen of fattening Japanese Black cattle, 10 bacterial taxa, showing >1% average relative abundance and >95% prevalence, irrespective of the dietary conditions and the fattening periods, were identified as the core rumen bacterial taxa, which accounted for approximately 80% of the rumen microbiota in Japanese Black cattle. Additionally, population dynamics of the core rumen bacterial taxa revealed two distinct patterns: Prevotella spp. and unclassified Bacteroidales decreased in the mid-fattening period, whereas unclassified Clostridiales, unclassified Ruminococcaceae, Ruminococcus spp., and unclassified Christensenellaceae increased during the same period. Therefore, the present study reports the wide distribution of the core rumen bacterial taxa in Japanese Black cattle, and the complementary nature of the population dynamics of these core taxa, which may ensure stable rumen fermentation during the fattening period.     

Differences in the behavior of the crossbred steers of different sire lines of Japanese Black cattle

We determined differences in the behavior of the progeny of two major sire lines of Japanese Black cattle by recording the behavior of 35 and 70 half-sib steers of sires from fast (FG) and slow (SG) growing lines, respectively. Two sire lines of steers were mixed and allocated to nine pens with 11–12 animals per pen. The proportion of steers lying was significantly (P < 0.001) higher in the SG line (43.4 ± 5.7% compared to 40.3 ± 6.0%). The proportion of time spent eating concentrate feed (FG: 12.1 ± 2.3%; SG: 11.4 ± 2.1%), drinking (FG: 0.8 ± 1.1%; SG: 0.4 ± 0.6%), licking the feed trough (FG: 0.4 ± 0.6%; SG: 0.2 ± 0.4%) and performing tongue-playing (FG: 3.1 ± 4.6%; SG: 1.0 ± 1.9%) was significantly higher in FG, whereas the proportion of time spent resting (FG: 41.5 ± 12.8%; SG: 43.7 ± 10.9%) and performing self-licking (FG: 1.7 ± 1.4%; SG: 2.1 ± 1.3%) was higher in SG (all P < 0.05). These results show progeny of the FG sire engaged in more active behaviors compared to the progeny of the SG sire line.     

Effect of probiotics on blood fatty acid metabolism during the late middle stage of fattening period in Japanese Black cattle

This study was conducted to investigate the effects of probiotics administration on fatty acid metabolism in Japanese Black cattle as per changes in blood fatty acid concentrations and blood biochemical tests. Eighteen clinically healthy Japanese Black female fattening cattle bred on the same fattening farm were randomly classified into the probiotics administration group (n=9) or the control group (n=9). In the probiotics administration group, 50 g of probiotics were started per animal per day at the age of 18 months, and the administration period was 2 months from the start date of the study. Blood was collected twice before starting the probiotics administration and at 2 months after starting the probiotics administration. In the probiotics administration group, palmitic, linoleic, arachidonic and α-linolenic acid tended to be higher at the end of the administration compared with those before probiotics administration. Additionally, as a result of multiple comparison test, monounsaturated fatty acids at Post was significantly higher, and the ω6 / ω3 ratio was significantly lower than in the control group. Vitamin A, E and albumin were significantly higher at the end of the administration than in the control group. In this study that administering probiotics to Japanese Black cattle in the late middle stage of fattening period did not have a significant effect on fatty acid metabolism during feed digestion and absorption, but suggested that may alter some blood fatty acids concentrations.     

Genome‐wide association study for fatty acid composition in Japanese Black cattle

Fatty acid composition is one of the important traits in beef. The aim of this study was to identify candidate genomic regions for fatty acid composition by genome‐wide association study with 50 K single nucleotide polymorphism (SNP) array in Japanese Black cattle. A total of 461 individuals and 40 657 SNPs were used in this study. We applied genome‐wide rapid association using mixed model and regression (GRAMMAR) and genomic control approaches to estimate the associations between genotypes and fatty acid composition. In addition, two SNPs in fatty acid synthase (FASN) (T1952A) and stearoyl‐CoA desaturase (SCD) (V293A) genes were also genotyped. Association analysis revealed that 30 significant SNPs for several fatty acids (C14:0, C14:1, C16:1 and C18:1) were located in the BTA19 FASN gene located within this region but the FASN mutation had no significant effect on any traits. We also detected one significant SNP for C18:1 on BTA23 and two SNPs for C16:0 on BTA25. The region around 17 Mb on BTA26 harbored two significant SNPs for C14:1 and SNP in SCD in this region showed the strongest association with C14:1. This study demonstrated novel candidate regions in BTA19, 23 and 25 for fatty acid composition.     

Identification of leptin gene polymorphisms associated with carcass traits and fatty acid composition in Japanese Black cattle

Previous studies have indicated that some leptin gene polymorphisms were associated with economically important traits in cattle breeds. However, polymorphisms in the leptin gene have not been reported thus far in Japanese Black cattle. Here, we aimed to identify the leptin gene polymorphisms which are associated with carcass traits and fatty acid composition in Japanese Black cattle. We sequenced the full‐length coding sequence of leptin gene for eight Japanese Black cattle. Sequence comparison revealed eight single nucleotide polymorphisms (SNPs). Three of these were predicted to cause amino acid substitutions: Y7F, R25C and A80V. Then, we genotyped these SNPs in two populations (JB1 with 560 animals and JB2 with 450 animals) and investigated the effects on the traits. Y7F in JB1 and A80V in JB2 were excluded from statistical analysis because the minor allele frequencies were low (< 0.1). Association analysis revealed that Y7F had a significant effect on the dressed carcass weight in JB2; R25C had a significant effect on C18:0 and C14:1 in JB1 and JB2, respectively; and A80V had a significant effect on C16:0, C16:1, C18:1, monounsaturated fatty acid and saturated fatty acid in JB1. The results suggested that these SNPs could be used as an effective marker for the improvement of Japanese Black cattle.     

Bos indicus type of growth hormone receptor gene is retained in Japanese Black cattle

The growth hormone receptor (GHR) gene is responsible for growth and carcass traits, and polymorphisms associated with the variation of meat production are thought to occur in the liver‐specific promoter of the GHR gene in cattle. The aim of this study was to analyse the structure of the liver‐specific promoter of GHR in Japanese Black cattle, as the relationship between GHR polymorphism and meat production is poorly understood in this breed. Typically in European cattle, the LINE‐1 element, a family of retrotransposons, is inserted in the liver‐specific promoter. However, a short GHR promoter without the LINE‐1 sequence was found in the Japanese Black breed as in Bos indicus cattle. The frequency of the short allele was approximately 60%. In addition, 24 of 29 Holstein/Japanese Black crosses carried the short allele from their sire. The present result suggests that the short allele for GHR may be a candidate marker for improving meat production of Japanese Black cattle.     

Effects of genetic and environmental factors on muscle glycogen content in Japanese Black cattle

Monosaccharides such as glucose contribute to the development of meat flavor upon heating via the Maillard reaction; therefore, monosaccharide content is related to beef palatability. Here, we analyzed the effects of genetic and environmental factors on the content of glycogen, one of the precursors of monosaccharides, in the muscles of 958 fattened Japanese Black cattle from Yamagata Prefecture. Analysis of variance showed that muscle glycogen content was affected by the farm and postmortem periods, but not by sex, slaughter age, slaughter month or number of days detained at the slaughter yard. Additionally, consumption of digestible brown rice feed elevated muscle glycogen levels. Glycogen heritability was estimated to be 0.34, and genetic correlations between glycogen and carcass weight (CW) or beef marbling standard (BMS) were weak. The predicted breeding values varied among paternal lines. These results demonstrated that genetic factors might improve muscle glycogen content and therefore beef palatability, but do not influence CW or BMS.     

固始鸡免疫器官内T淋巴细胞亚群动态变化

应用流式细胞仪结合免疫荧光抗体技术对固始鸡和固始鸡胚胎免疫器官内T淋巴细胞亚群的动态变化和发育分化规律进行了研究.结果发现,(1)胚胎时期,胸腺内T淋巴细胞发育分化低,表现为成熟的T淋巴细胞(CD3~+)和不成熟的胸腺细胞均较少;出壳后,固始鸡胸腺内成熟T淋巴细胞增多,占胸腺细胞的30%左右,但大部分仍是未成熟的胸腺细胞.(2)脾脏内T淋巴细胞为成熟的T淋巴细胞(CD3~+),占淋巴细胞的60%左右.胚胎时期脾脏内CD3~+水平低,出壳后2周已达脾脏内正常水平(60%左右).(3)法氏囊内有少量的成熟T淋巴细胞(10%左右).结果表明,T淋巴细胞在固始鸡胸腺内增殖分化和成熟,脾脏和法氏囊内的成熟T淋巴细胞均来源于胸腺.     

Three cases of idiopathic eosinophilic enteritis with chronic obstinate diarrhea in Japanese Black fattening cattle

Eosinophilic enteritis (EOE) is a type of inflammatory bowel disease and is characterized clinically by chronic obstinate diarrhea. Three Japanese Black (JB) fattening cattle (2 males and 1 female) on different cattle farms presented with chronic episodic diarrhea without fever or dehydration. Soft reddish spherical carneous tissues (1−3 cm) were occasionally excreted within the diarrheic feces. Administration of antibiotics, antidiarrheal drugs and vermicides had no therapeutic effect, but dexamethasone improved the fecal characteristics. The symptoms persisted until the animals were slaughtered at 27–30 months of age. Histopathological examination of the intestines revealed marked eosinophilic infiltration in the lamina propria and submucosa. From these findings, we diagnosed these cattle as the first cases of EOE in JB cattle.     

The assessment of genetic diversity within and among the eight subpopulations of Japanese Black cattle using 52 microsatellite markers

Japanese Black cattle are at risk for genetic homogeneity due to intensive use of a few sires. Therefore, assessment of the actual genetic diversity of this breed is important for future breeding plans. In the present study, we investigated the genetic diversity within and among eight subpopulations of Japanese Black cattle using 52 microsatellite markers. The parameters for genetic diversity of Japanese Black cattle were comparable to those of other cattle breeds, suggesting that the relatively high genetic diversity of the breed. However, upon comparison among the eight subpopulations, the Hyogo subpopulation showed markedly low genetic diversity. The results of the pairwise F_ST values, phylogenetic network and structure analysis indicated that the Hyogo population has remarkably high level of genetic differentiation from other populations, while Yamagata, Niigata, Hiroshima and Kagawa populations have low levels of genetic differentiation. Furthermore, multidimensional scaling plots indicated that individuals in some subpopulations were separated from individuals in the other subpopulations. We conclude that while the overall genetic diversity of Japanese Black cattle is still maintained at a relatively high level, that of a particular subpopulation is significantly reduced, and therefore the effective population size of the breed needs to be controlled by correct mating strategies.     

Estimation of genetic parameters for carcass defects of Japanese Black cattle in Kagoshima

Cattle exhibit a range of carcass defects, including blood splash (BLS), intramuscular edema (INE), muscle steatosis (MUS), bruising (BR), trim loss (TRL) and others (OTH). These defects lower the carcass value and can result in significant economic loss to producers. We estimated the incidence, relationship with inbreeding coefficients and genetic parameters of carcass defects in Japanese Black cattle using 561 619 carcass records from Kagoshima, Japan during April 1988 through March 2011. The defect incidence ranged from 0.22% for TRL to 5.73% for BR. The incidence of MUS and BR increased from 1.21% to 6.57% and from 1.06% to 9.31%, respectively. The incidence of INE peaked at 7.44% in 1999 and decreased thereafter. We observed a positive linear relationship between the defect incidence and the inbreeding coefficients in MUS, BR and TRL (P < 0.01). The heritabilities estimated by univariate animal model with Gibbs sampling for BLS, INE, MUS, BR and TRL were 0.24, 0.06, 0.18, 0.05 and 0.02, respectively. The contribution of farm variance to phenotypic variance was negligible (0.01 to 0.04). Significant genetic correlations of TRL were estimated with MUS (0.63) and BR (0.63). Our results suggest that genetic factors contribute to the incidence of BLS and MUS.     

Performance of using opposing homozygotes for paternity testing in Japanese Black cattle

Genome-wide single nucleotide polymorphism (SNP) markers in Japanese Black cattle enable genomic prediction and verifying parent–offspring relationships. We assessed the performance of opposing homozygotes (OH) for paternity testing in Japanese Black cattle, using SNP genotype information of 50 sires and 3,420 fattened animals, 1,945 of which were fathered by the 50 genotyped sires. The number of OH was counted for each sire–progeny pair in 28,764 SNPs with minor allele frequencies of ≥0.05 in this population. Across all pairs of animals, the number of OH tended to increase as the pedigree-based coefficient of relationship decreased. With a threshold of 288 (1% of SNPs) for paternity testing, most sire–progeny pairs were detected as true relationships. The frequency of Mendelian inconsistencies was 2.4%, reflecting the high accuracy of pedigree information in Japanese Black cattle population. The results indicate the utility of OH for paternity testing in Japanese Black cattle.     

Genetic relationships between temperament of calves at auction and carcass traits in Japanese Black cattle

Correlations of calves’ temperament with carcass traits were estimated to clarify the genetic relationships between them in Japanese Black cattle. The temperament records for 3128 calves during auction at a calf market were scored on a scale of 1 (calm) to 5 (nervous) as temperament score (TS ), and the TS were divided into two groups (TSG ): TS 1 and 2 comprised TSG 1, and 3 to 5 constituted TSG 2. Carcass data were obtained from 33 552 fattened cattle. A threshold animal model was used for analyzing the underlying liability for TSG , whereas a linear one was used for TS and carcass traits. The heritability estimates for TS and TSG were 0.12 and 0.11, respectively. On the other hand, moderate to high heritability estimates were obtained for carcass traits (0.40 to 0.68). The temperament scores were negatively correlated with carcass weight, rib thickness and subcutaneous fat thickness (?0.13 to ?0.59). In contrast, weak to moderate positive correlations were found between the temperament scores and rib eye area or yield estimate (0.16 to 0.45). The temperament scores and beef marbling score had no correlation. These results showed that it is possible to improve temperament and carcass traits simultaneously.     

Genomic imprinting variances of beef carcass traits and physiochemical characteristics in Japanese Black cattle

The objective of this study was to estimate variance components related to imprinting for carcass traits and physiochemical characteristics in Japanese Black cattle. The carcass records obtained from 4,220 Japanese Black feedlot cattle included carcass weight (CW), rib eye area (REA), rib thickness, subcutaneous fat thickness, and beef marbling score (BMS), and the physiochemical characteristics were fat, moisture, glycogen per proportion of moisture content, oleic acid, and monounsaturated fatty acids (MUFA). To detect gametic effects, an imprinting model was fitted. High additive heritabilities were estimated for all traits (from 0.516 for glycogen to 0.853 for fat) and were reduced in Mendelian heritability. The range of the differences was from 0.002 (CW) to 0.331 (fat and moisture), and the reductions were due to their imprinting variances. The ratio of the imprinting variance to the total additive genetic variance for REA (0.374), BMS (0.291), fat (0.387), moisture (0.388), and MUFA (0.337) were large (p < 0.05). These imprinting variances were due to the maternal contribution and suggested the existence of maternally expressed genomic imprinting effects on the traits in Japanese Black cattle. Therefore, maternal gametic effects should be considered in breeding programs for Japanese Black cattle.     

Comparison of the peripheral blood leukocyte population between Japanese Black and Holstein calves

Japanese black (JB) calves have greater susceptibility to infectious diseases compared to Holstein (Hol) calves. In order to clarify the differences in cellular immune status between JB and Hol calves, the leukocyte population and lymphocyte proliferative ability were analyzed. In total 200 healthy calves, 1 day to 14 weeks of age, were examined: 105 JB and 95 Hol calves. Lower numbers in peripheral blood and percentage in peripheral blood mononuclear cells of CD3⁺TcR1‐N12⁺ T cells and major histocompatibility complex class‐II⁺CD14^‐ B cells were observed in the JB compared to the Hol. The percentage of TcR1‐N12⁺CD25⁺ T cell in the JB was significantly lower than that of the Hol at 4–6, and 8–10 weeks. Interleukin (IL)‐2 sensitivity in the JB was lower than that in the Hol, and significant differences were observed in age groups of 6–8 weeks and 10–14 weeks. These findings indicated that the lower numbers of γδ T cells and B cells in the JB compared to the Hol might be associated with the specificity of the immune systems in JB calves.     

Relationship between serum leptin concentrations and the marbling scores in Japanese Black Cattle

The objective of this study was to investigate the relationship between marbling and serum leptin concentrations. First, we compared serum leptin concentrations between Japanese Black cattle and Holstein calves, and found that the mean serum concentration was significantly higher (P < 0.05) in Japanese Black cattle (2.67 ± 0.41 ng/mL, n = 13) than in Holstein calves (1.79 ± 0.43 ng/mL, n = 5). We then examined the effects of roughage intake or vitamin A on the serum leptin concentrations in Japanese Black cattle. Although the leptin concentrations for the high roughage group were significantly higher than those for the low roughage group at 4, 7 and 20 months old, there was no significant difference in the carcass characteristics of the two groups. In addition, although there was a significant difference in the marbling score between the high vitamin A group and low vitamin A group, there was no significant difference in serum leptin concentrations between the groups. These results suggest that although serum leptin concentrations were not significantly correlated to the marbling scores they may be related to the genetic advantage for marbling in Japanese Black cattle.     

Genetic parameters for gestation length and the relationship with birth weight and carcass traits in Japanese Black cattle

The objectives of this study were to estimate genetic parameters for gestation length (GL), including estimation of maternal effects, and to investigate the genetic relationships of GL with birth weight and carcass traits in a Japanese Black cattle population. The original data comprised 34 775 records of animals born from October 1999 to August 2003. Two different models were used to analyze the data for GL. The first model (M1) included direct genetic effect of the calf and maternal genetic effect as random effects. The second model (M2) treated GL as a trait of the dam and included direct genetic effects only. M1 was used in bi-variate analysis. The direct and maternal heritabilities for GL estimated from M1 were 0.53 and 0.14, respectively. This result shows that GL is moderately inherited and can be controlled genetically. The direct × maternal genetic correlation for GL was −0.73. Direct genetic correlations of GL with carcass traits were close to zero. However, genetic correlation of maternal GL with carcass weight was moderate (0.25).     

Nucleotide sequence of myostatin gene and its developmental expression in skeletal muscles of Japanese Black cattle

Myostatin, a member of the transforming growth factor‐β superfamily, is a well known negative regulator of skeletal muscle growth. In the present study, the 6660 bp nucleotide sequence of the myostatin gene in Japanese Black cattle (JBC), including the entire coding region of 1128 bp, was determined. The amino acid sequence deduced from the nucleotide sequence of JBC was well conserved with its sequence of other cattle, although it was found that an Α→G transition at nucleotide position 641 results in the substitution of asparagine by serine at amino acid position 214. In order to examine the expression pattern of the myostatin gene in the skeletal muscles of JBC, its expression in three skeletal muscles, Semitendinosus (ST) muscle, Biceps femoris muscle and Longissimus lumborum muscle, of fetal and calf stages was analyzed by real time polymerase chain reaction. The highest level of the myostatin expression was observed in the fetal stage. In calf stages the highest expression was observed in ST muscle compared with the other two muscles. These results suggest that a higher expression of myostatin gene, especially in the fetal stage and in ST muscle during calf stages, is involved in the arrest in skeletal muscle growth and that its functional domains and genomic structure in JBC are well conserved with those in other mammals.