Detecting endotoxin activity in bovine serum using an automated testing system

The aim of the present study was to compare the ability of the commercially available portable test system (PTS^TM) to detect endotoxin activity in bovine serum, with that of the traditional LAL-kinetic turbidimetric (KT) and chromogenic (KC) assays. Prior to testing, serum samples, which were obtained from endotoxin-challenged cattle, were diluted 1:20 in endotoxin-free water and heated to 80°C for 10 min. The performance of the PTS^TM was not significantly different from that of the traditional LAL-based assays. The results using PTS^TM correlated with those using KT (r²=0.963, P<0.001) or KC assays (r²=0.982, P<0.001). Based on these findings, the PTS^TM could be applied as a simplified system to assess endotoxin activity in bovine serum.     

转移因子注射液中细菌内毒素检测方法研究

为检测转移因子(TF)注射液中细菌内毒素,本研究采用鲎试剂与细菌内毒素产生凝集反应的凝胶法对国内1个生物制品厂的TF注射液样品进行了检测,实验结果表明当TF注射液稀释至0.025 mg/mL时,对鲎试剂的凝集反应无干扰作用.因此,该方法适用于TF注射液中细菌内毒素检测,并可以用于该项生物制品的质量监控.     

Evaluation of a Cow-Side Test for Detection of Gram-Negative Bacteria in Milk from Cows with Mastitis

Waage, S., P. Jonsson and A. Franklin: Evaluation of a cow-side test for detection of Gram-negative bacteria in milk from cows with mastitis. Acta vet. scand. 1994, 207-211.– A modified Limulus amebocyte lysate (LAL) cow-side test was evaluated under field conditions. The principle of the test is to visualize reactions between test components and endotoxin from the cell wall of Gram-negative bacteria. The practical purpose is to detect such bacteria in mastitic milk. Secretions from 789 udder quarters with clinical mastitis were examined by the LAL-test. Parallel quarter milk samples were sent to a mastitis laboratory for microbiological examination. Eleven veterinary surgeons in three veterinary districts in Norway performed the field investigations. Results of the LAL-test and culture agreed in 93% of all milk samples tested, agreement measured by kappa being 0.63. The sensitivity of the test in detecting Gram-negative bacteria was 63%, while the specificity was 97%. The predictive value of a positive test result was 70%, the figure being somewhat higher (75%) when the material was limited to milk samples without antibiotic residues. The predictive value of a negative test result was 95%. The LAL-test is considered to constitute a valuable cow-side test for the veterinary practitioner, aiding the selection of antibacterial drug of choice for the initial treatment of clinical mastitis.     

影响江苏某牛场荷斯坦牛日产奶量和乳成分因素分析

[目的]探究江苏某牛场荷斯坦牛日产奶量和乳成分的影响因素。[方法]试验采集了该规模化牛场2018—2020年139 703条测定数据,并利用多因素方差模型对其进行系统的分析。[结果]不同胎次、测定季节、产犊季节、泌乳月对荷斯坦牛日产奶量、乳脂率、乳蛋白率、体细胞评分、乳尿素氮均有极显著影响(P<0.01)。产奶量与乳脂率、乳蛋白率、体细胞评分均存在极显著负相关。[结论]综上结果,在生产中,应结合胎次、季节、产犊时间、泌乳等多种因素,灵活调整牛群结构、生产规划和饲养管理,以实现提高产奶量和乳品质的目的。     

Use of a test day model for dairy goat milk yield across lactations in Germany.

Breeding value prediction for dairy goats in Germany is still based on herd mate comparison within breeding society. The objective of this study was to estimate genetic parameters for milk yield based on a test day model. For the analysis 35,308, 30,551 and 23,640 test day records from lactations 1, 2 and 3 from 5079, 4118 and 3132 animals, respectively, were used. The data between 1987 and 2003 were obtained from six German breeding societies. The multiple trait (lactations 1, 2 and 3) repeatability model (RPT) included the fixed effects of breeding society-breed-herd-year, litter size, lambing season, and days in milk of third-order Legendre polynomials nested within herd-year, and the random effects of animal additive and permanent environment. The three-trait random regression model (RR) also included the random regressions based on second-order Legendre polynomials for animal additive and permanent environmental effects. Heritability estimates in RPT were 0.27 +/- 0.02, 0.20 +/- 0.02 and 0.37 +/- 0.02 for the first, second and third lactation, respectively. The genetic correlation between the first and second lactation was 0.69, between the second and third lactation 0.79, and between the first and third lactation 0.45. Heritability estimates from the RR in the first and second lactations decreased from the beginning to the end of the lactation, with average values of 0.28 and 0.27, respectively. Estimates in the third lactation showed a maximum in the middle of lactation, averaging 0.37. Genetic correlations between the first and second lactation averaged 0.64, between the second and third lactation 0.72, and between the first and third lactation 0.46. Despite the small data set and restricted relationship structure the estimates were reasonable with the exception of estimates from the third lactation, which seemed inflated. RR could be used for genetic evaluation of dairy goats in Germany.     

Effects of long-term administration of recombinant bovine somatotropin on the plasminogen–plasmin system and milk composition of crossbred Holstein cattle

Experiments were performed to examine the effects of long‐term treatment with recombinant bovine somatotropin (rbST) on milk plasmin–plasminogen and milk composition by one injection of 500 mg of rbST in every 14 days throughout lactation in crossbred Holstein cattle. The animals receiving rbST gave a greater milk yield and rate of blood flow to the udder during early lactation than the control group. The milk lactose concentration remained constant while the milk protein concentration increased as lactation advanced in both groups. The level of milk fat in rbST‐treated animals was significantly greater than in controls in the early lactation period (P < 0.05). The milk sodium and chloride concentrations of the rbST‐treated animals significantly decreased in early lactation as compared with the control animals. The sodium : potassium ratio of the rbST‐treated animals was significantly lower than those of control animals in the early lactation (P < 0.05) and it markedly increased in late lactation. As lactation advanced, the concentration of plasmin in the milk gradually increased, while the milk plasminogen concentration significantly increased in both groups. The plasminogen : plasmin ratio decreased in the control animals while it increased in the rbST‐treated animals as lactation advanced. These findings demonstrate that rbST is involved the activity of the plasmin–plasminogen system but is not involved in maintaining tissue integrity in the mammary gland during late lactation in crossbred dairy cattle.     

Storage strategies and processing of bovine milk samples for pregnancy-associated glycoproteins detection test

Three experiments were performed to test if the time and storage conditions of milk samples, the preheating of samples in a water bath, as well as the carryover effect in laboratory analysis equipment could affect the pregnancy-associated glycoproteins (PAG) levels, and consequently the results of a pregnancy test. The pregnancy test used in both experiments uses the enzyme-linked immunosorbent assay method to measure the concentrations of PAG and classify the samples as pregnant, nonpregnant or suspicious. As a result, PAG levels showed no variation when the samples were analyzed up to 9 days after collection, whether stored in ambient temperature or refrigerated. The preheating of the samples in a water bath and prior to the analysis of SCC and composition also did not affect the levels of PAG, allowing the same sample used in the quality analysis to be used for the pregnancy test.     

Real‐time quantitative PCR‐based detection of Coxiella burnetii in unpasteurized cow's milk sold for human consumption

Coxiella burnetii is a zoonotic pathogen with a worldwide distribution that is responsible for Q fever in humans. It is a highly infectious bacterium that can be transmitted from cattle to humans through the consumption of unpasteurized milk. We report the molecular identification of C. burnetii in raw cow's milk being sold directly for human consumption in Brazil without official inspection or pasteurization. One hundred and twelve samples of raw milk were analysed by real‐time quantitative PCR (qPCR), and C. burnetii was detected in 3.57% (4/112) of the samples at a concentration ranging from 125 to 404 bacteria per millilitre. The identification of this zoonotic pathogen in raw milk sold directly for human consumption is a public health concern since C. burnetii can be transmitted through the oral route. This result indicates that health education and other preventive measures should be officially implemented in Brazil to prevent the spread of infection. To our knowledge, this is the first qPCR‐based detection of C. burnetii in raw milk samples from cows sold in Brazil that do not undergo official inspection or pasteurization.     

Evaluation of a portable device for assessment of motility in stallion semen

In horse breeding, quality assessment of semen before insemination is often requested. Non‐laboratory‐based techniques for objective analysis of sperm motility are thus of interest. The aim of this study was evaluating a portable device for semen analysis (Ongo sperm test) and its comparison with computer‐assisted semen analysis (CASA). Semen was collected from 10 stallions, diluted to 100, 50 and 25 × 10⁶ sperm/ml and analysed for total (TM) and progressive motility (PM). The final sperm concentration influenced total motility analysed by Ongo (p < 0.05) which was higher at 100 × 10⁶ sperm/ml when compared to 25 × 10⁶ sperm/ml (p < 0.05) but not when compared to 50 × 10⁶ sperm/ml (n.s.). Sperm concentration did not influence total motility when assessed by SpermVision (n.s.). Agreement between methods was evaluated by correlation analysis and Bland–Altman plot. Intra‐assay variation of Ongo was 5.2% ± 3.0 for TM and 6.9% ± 3.4 for PM. Correlation between Ongo and CASA was r = 0.79, 0.88 and 0.83 for 100, 50 and 25 × 10⁶ sperm/ml for TM, and r = 0.87, 0.89 and 0.87 for PM, respectively (all p < 0.001). At the 100 and 25 mio/ml dilutions, the difference between the two systems deviated significantly from 0, while no such bias existed at the 50 mio/ml dilution (TM Ongo 85.0%, CASA 82.3%; PM Ongo 64.1%, CASA 66.1%). The 95% confidence interval was 19.9%, 18.9% and 19.2% ± mean for TM and 20.7%, 17.4% and 20.3% ± mean for 100, 50 and 25 × 10⁶ sperm/ml, respectively. In conclusion, Ongo sperm test sperm motility data were strongly correlated with data obtained by CASA. In addition, at a concentration of 50 × 10⁶ sperm/ml values measured with both systems were close to identical. At this concentration, which is recommended in equine AI, Ongo and CASA can be used interchangeably.     

牛奶和血清中胰蛋白酶活性的体外检测试验

为了给胰蛋白酶为主要成分的药物在奶牛体内的药代动力学提供合适的研究方法,以体外添加胰蛋白酶的奶牛血清和牛奶为研究对象,优化商品化的胰蛋白酶活性检测试剂盒,采用紫外分光光度法检测奶牛血清和牛奶中胰蛋白酶的活性。结果显示:0~0.4 nmol·m L-1范围内,胰蛋白酶的吸光度值与浓度呈现良好的线性关系,回归方程y=0.023x-0.003 8,r2=0.999 6;以信噪比S/N=3为标准,在牛奶中最低检测限浓度为25 ng·m L-1,平均回收率为84.3%,3 d中日内变异系数分别为13.9%、10.2%、11.8%,日间变异系数为11.0%;在血清中最低检测限浓度为15 ng·m L-1,平均回收率为85.6%,3 d中日内变异系数分别为13.7%、13.2%、8.1%,日间变异系数为13.8%。结果表明:该方法检测牛奶和血清中的酶活准确率高,在试剂盒可检测的浓度范围内受牛奶和血清中的蛋白成分影响很小,且回收率和精密度也都达到了兽用药物药代动力学试验指导原则的要求,具有良好的准确性和重现性。     

Evaluation of a monitoring system for assessing animal health in dairy cattle

A monitoring system to assess animal health in dairy farms was developed and applied on 44 farms in an epidemiological study. The objective of the study was to evaluate the applicability of the system for long term monitoring of health data. For this purpose, the quality of the collected data and the acceptance of the system by farmers and veterinarians were investigated. Farmers recorded all health events using a form. Collected data was entered into a central database on a monthly basis. The recording of health data was easy to do and did not take a lot of time. Most of the farmers (89%) used less than 20 minutes per week for the recording. This time need was independent of the herd size. 66% of the farmers found it useful to record health data, especially for improving their overview on health of each single cow. Treatments were reported in 81% of the cases, inseminations and calving in 80% of the cases. All events were reported correctly and precise.     

Development of novel SNP system for individual and pedigree control in a Japanese Black cattle population using whole‐genome genotyping assay

Individual identification and parentage analysis using DNA markers are essential for assuring food identity and managing livestock population. The objective of this study was to develop a single nucleotide polymorphism (SNP) panel system for individual effective identification and parentage testing in a Japanese Black cattle population using BovineSNP50 BeadChip. On the basis of SNP frequencies, 60 unlinked informative SNPs were finally selected as candidate markers. The allelic frequencies for each SNP were estimated using additional individuals by PCR‐RFLP (restriction fragment length polymorphism). A total of 87 SNP markers added in conjunction with previously developed 27 SNPs were evaluated to assess the utility of the test. The estimated identity power was 2.01 × 10^?34. Parentage exclusion probabilities, when both suspected parents' genotypes were known and when only one suspected parent was genotyped, were estimated as 0.99999997 and 0.99998010, respectively. This developed SNP panel was quite powerful and could successfully exclude false sires with a probability of >0.9999 even if the dam's genotype information was not obtained. The SNP system would contribute to management of the beef industry in Japan.     

Improved milk production through PG–PL system by provision of in‐house shelter management in lactating Murrah buffaloes during winter season

Comprehensive information on the role of β‐casein and plasminogen–plasmin (PG–PL) system in milk secretion of Murrah buffaloes during winter season is lacking, although effects of cold stress can be ameliorated to an extent by altering microclimate at farm level. Hence, this study was aimed to determine the changes in productivity along with PG–PL system of milk, plasma hormones and metabolites of buffaloes during winter (December–January) season under two different management systems. Average minimum temperature and wind chill index during this season were 7.02 and 12.74 °C respectively. Buffaloes were divided in two groups of six animals each: control and treatment, where treatment group animals were placed in‐house with floor bedding of paddy straw and the control group animals in loose housing system without straw bedding. Physiological responses were recorded, and milk and blood samples were collected at weekly intervals for six‐week experimental period. Under in‐house management system, buffaloes experienced better comfort by alleviating environmental stress as their physiological responses such as respiration rate and pulse rate were significantly reduced (p < 0.01) as compared to the control, which subsequently resulted higher milk yield by 9.92% (p < 0.05). Analysis of milk samples revealed higher concentration of plasminogen (10.6 vs. 8.05 μg/ml; p < 0.01) and β‐casein (p < 0.05), and lower plasmin level (0.299 vs. 0.321 μg/ml; p < 0.05) in buffaloes under treatment group. It was also found that plasma cortisol, glucose and non‐esterified fatty acids levels were higher (p < 0.01) in control group as compared to the treatment animals by 13.6%, 8.14% and 12.6% respectively. However, milk composition, growth hormone, epinephrine and norepinephrine level in plasma were similar in both the groups. Hence, it may be concluded that provision of in‐house shelter management with floor bedding of paddy straw during winter was effective to minimize environmental stress and improved milk production through manipulation of PG–PL system in buffaloes.     

Evaluation of a rapid coliform detection kit from clinical mastitis milk using colloidal gold nanoparticle–based immunochromatographic strips

The accurate identification of mastitis‐causing bacteria assists in effective management by both dairy farmers and veterinarians and can be used to implement the selective use of antimicrobials for treatment. The purpose of this study was to evaluate the ability of our developed anti–ribosomal protein-L7/L12 antibody–coated immunochromatographic strip (ICS) test to detect coliforms in milk by comparing the results with the bacteriological culture method. We investigated the performance of the ICS test as compared with the bacteriological culture method using 308 milk samples from clinical bovine mastitis. First, to determine the optimal ICS test cutoff point for detecting coliform mastitis, we developed a receiver-operating characteristic curve. The result showed that the cutoff point was at 0.5 of our index. The sensitivity, specificity, positive predictive value (PPV), and negative predictive value of the ICS test were 81.3%, 84.8%, 69.2%, and 91.54%, respectively. As the clinical signs increased in severity, the F-measure, a weighted harmonic mean of the sensitivity and overall PPV performance, increased. Because it is especially important to treat clinical mastitis appropriately in the early stages of detection, the ICS test, which can be used by both dairy farmers and veterinarians on dairy farms, is considered to be a useful tool for detecting coliform mastitis, which often presents with severe signs.     

Development of genetic evaluation for milk production traits of Holsteins in Japan

The procedure used for the genetic evaluation of dairy cattle in Japan has developed from a lactation sire–MGS model to a multiple‐lactation random regression test‐day animal model. Genetic evaluation of Holstein bulls in Japan began in 1989 with the use of field‐style progeny testing; dairy herd improvement program data from all over Japan were used, along with a sire and maternal grandsire model. In 1993, an animal model was introduced to estimate breeding values for yield and type traits. A random regression test‐day model was first applied in 2010. In the business of breeding dairy cattle, it is very important to users that estimated breeding values are reliable and stable among subsequent routine evaluations. With experience in the genetic evaluation of dairy cattle in Japan, Japanese researchers have found ways to improve the stability of estimated breeding values. These modifications involve changes in data editing, development of evaluation models, changes to the structures of unknown‐parent groups, awareness of the problems of predicting lactation yield from partial test‐day records, and adjustment for heterogeneity within herd variances. Here, I introduce developments in, and our experiences with, the genetic evaluation of yield traits of Holstein cattle in Japan.     

Evaluation of enzyme-linked immunosorbent assay using milk samples as a potential screening test of bovine tuberculosis of dairy cows in Korea

An enzyme-linked immunosorbent assay (ELISA) using bulk tank milk samples was evaluated as a screening test for bovine tuberculosis (TB), a contagious chronic disease of cattle. An ELISA with MPB70, a major antigen of Mycobacterium bovis was performed using paired sets of milk and sera samples from 33 tuberculin-positive and 43 tuberculin-negative cattle. Anti-MPB70 antibodies were detected in milk samples and there was a significant correlation between seroreactivities of milk and sera samples (R² = 0.83). Using the tuberculin skin test as the reference test, the sensitivities of ELISA using milk and sera samples were 87.8% and 81.8%, respectively, and the specificities were 97.7% and 100%, respectively.In the screening test using bulk tank milk samples from 931 dairy herds in Whasung, Gyeonggi-do, Korea, the positive rate for anti-MPB70 antibody was 4.5% (42/931) and the tuberculin-positive rate was 2.8% (26/931). Individual milk samples (n = 253) were collected from randomly selected 8 problematic and 3 negative herds (positive and negative in the screening test by MPB70 ELISA using bulk tank milk samples, respectively) and tested by MPB70 milk ELISA. In the problematic herds, positive rates were 10.5% (20/190) for anti-MPB70 antibodies in milk ELISA and 2.1% (4/190) in the tuberculin skin test. More than one dairy cows were positive by milk ELISA among the problematic herds, and all tuberculin-positive dairy cows were positive in the milk ELISA. Further, no positive cows were detected in negative herds both by milk ELISA and tuberculin skin test. These results suggest that an ELISA, using bulk tank milk samples, might be a potential efficient screening test for bovine TB of dairy cows.     

Evaluation of a Rapid Method for the Determination of Urea in Cow’S Milk

A rapid method for the determination of urea in milk was evaluated. The method employed was a urease/glutamate dehydrogenase procedure where the decrease in absorbante of NADH is monitored spectrophotometrically at 340 nm. The repeatability expressed as a coefficient of variation, was 1.4 % and 0.8 % for whole milk and skim milk, respectively. The accuracy of the method for urea determinations in whole milk was satisfactory (coefficient of variation, 2.6 %). Adding the preservative bronopol (0.02%) to the milk or freezing of the milk samples for one week did not alter urea levels significantly (P > 0.05). The storage of preserved whole milk in a refrigerator for 14 days, however, increased milk urea levels measured by 3.1 % (P < 0.001).It was concluded that the method evaluated is simple, rapid (40–50 samples/h) and yields reliable results also when whole milk is used. Since the determination of milk urea may be useful in assessing the feeding of energy and protein, the method is suitable for integration in the existing milk recording system in Sweden.     

Feed handling of lactating crossbred cows maintained in a semi‐arid region during the hot season: physiological parameters,ingestive behavior and performance

The effects of time of feed delivery (14.00 hours; 14.00 and 20.00 hours; 16.00 and 18.00 hours) on the physiological parameters, ingestive behavior, nutrient intake and production of lactating cows maintained in a semi‐arid region during the hot season were evaluated. Regardless of treatment, all animals received the first feeding supply at 06.00 hours. Eight cows with an average body weight of 600 kg, average milk yield of 20 kg/day and 80 days in milk were utilized. The rectal temperature, respiratory rate and sweating rate were not affected (P > 0.05), with average values of 38.5°C, 53.8 movements/min and 104 g/m²/h respectively. There was no effect (P > 0.05) on the eating time (314 min/day), ruminating time (564 min/day), drinking time (61 min/day) and idle time (502 min/day). Similarly, the intake of nutrients and performance of lactating cows were not affected (P > 0.05), with average dry matter intake of 19.8 kg/day, 4% fat‐corrected milk of 20.6 kg/day and milk fat concentration of 4.03 g/100 g. Since the behavior and performance have not altered, any times of feed delivery evaluated could be used to crossbred Holstein × Zebu cows maintained on a feedlot in semi‐arid regions during the hot season.     

Estimation of genetic parameters for milk and fertility traits within and between low,medium and high dairy production systems in Kenya to account for genotype-by-environment interaction

Dairy records from the Dairy Recording Service of Kenya were classified into low, medium and high production systems based on mean 305-day milk yield using the K-means clustering method. Milk and fertility records were then analysed to develop genetic evaluation systems accounting for genotype-by-environment interaction between the production systems. Data comprised 26,638 lactation yield, 3,505 fat yield, 9,235 age at first calving and 17,870 calving interval records from 12,631 cows which were descendants of 2,554 sires and 8,433 dams. An animal model was used to estimate variance components, genetic correlations and breeding values for the production systems. Variance components increased with production means, apart from genetic group variances, which decreased from the low to the high production system. Moderate heritabilities were estimated for milk traits (0.21–0.27) and fat traits (0.11–0.38). Low heritabilities were estimated for lactation length (0.04–0.10) and calving interval (0.03–0.06). Moderate heritabilities (0.25–0.26) were estimated for age at first calving, except under the high production system (0.05). Within production systems, lactation milk yield, 305-day milk yield and lactation length had high positive genetic correlations (0.52–0.96), while lactation milk yield and lactation length with age at first calving had negative genetic correlations. Milk yield and calving interval were positively correlated except under the low production system. The genetic correlations for lactation milk yield and 305-day milk yield between low and medium (0.48 ± 0.20 and 0.46 ± 0.21) and low and high production systems’ (0.74 ± 0.15 and 0.62 ± 0.17) were significantly lower than one. Milk yield in the low production system is, therefore, a genetically different trait. The low genetic correlations between the three production systems for most milk production and fertility traits suggested that sires should be selected based on progeny performance in the targeted production system.