| Heilongjiang Province is the largest high-end dairy production base in China.Dairy industry is the pillar industry of animal husbandry in Heilongjiang Province,and limb and hoof diseases is the second major problem after mastitis that affects the large-scale breeding of dairy cows.Limb and hoof diseases affects milk yield and reproductive performance,and even leads to the elimination of dairy cows,directly affecting the economic benefits of the pasture.The occurrence of limb and hoof disease is concealed and does not show clinical symptoms in the early stage.At present,the diagnosis method of cow limb and hoof disease is backward,and it is difficult to carry out early diagnosis.When the clinical symptoms are shown,the tissues have un dergone significant irreversible pathological changes,and the treatment is difficult to be eliminated,causing huge economic losses to animal husbandry.Therefore,in this study,the incidence of cow limb and hoof diseases in Heilongjiang Province was investigated and regional maps were drawn,and comprehensive diagnostic techniques for osteoarthritis and laminitis were studied.In the first part of this study,the incidence and causes of cow limb and hoof diseases in Heilongjiang province were investigated,and the incidence map was drawn at the same time.In view of the characteristics of many and complex factors of cow limb and hoof diseases in Heilongjiang province,the investigation was carried out from aspects of nutrition environment,feeding management and so on.The incidence map of deformed hooves,ulcers on the sole and tip of the hooves,white line disease,hoof dermatitis,laminitis,hoof rot disease and other limb and hoof diseases of dairy cows in Heilongjiang province was drawn to analyze the main factors causing limb and hoof disease in this area.The second part of this study was the study of comprehensive diagnostic techniques for osteoarthritis(OA,OARSI)and laminitis in dairy cows,aiming to establish rapid diagnostic methods and techniques for osteoarthritis and laminitis in dairy cows.In the screening test of sensitive biomarkers for bone and joint in dairy cows,the specific biomarkers were screened by using the rat model of osteoarthritis,and the disease model was established by anterior cruciate ligament amputation.The tibia and serum samples of the rats were collected at 0,2,4,6,8,10 weeks after the modeling.HE staining was perfor med on the tibia samples,and OARSI score was used to observe the histological changes of osteoarthritis.Cartilage degradation marker CTX-II,COMP,CS846,C2 C,GAG and PIICP in serum were detected by ELISA.The changes of concentration were analyzed to ana lyze the correlation between the changes of the concentration of OA markers and the degree of joint injury in rats,so as to screen the effective detection indicators of OA,and then the application test of the selected markers in cow OA was carried out.In cows,laminitis sensitive biomarker detection test,the low poly fructose infusion method is used to induce cows laminitis model,induced at 6 h intervals during continuous monitoring clinical indicators of cows,disease development,to be euthanized aft er 72 h,take the hoof tissue HE and Safranin O staining,collection building 0 h,6 h,12 h after 18 h.Serum samples were collected at 24 h,36 h,48 h,60 h and 72 h.Serum expressions of MMP 9 and TIMP1 were detected by gelatin enzyme spectrum and rever se gelatin enzyme spectrum.This survey was conducted from August 2018 to November 2020,with a total of 29,050 cows from 27 dairy farms covering five regions in Heilongjiang Province.The survey results showed that the four most prevalent limb and hoof diseases were hoof dermatitis(16.4%)and deformed foot(4.2%),respectively.Ulcers on the sole and tip of the hoof(3.5%)and white line disease(1.2%).By analyzing regional map,the cow limb hoof disease incidence from low to high followed by Daqing,Qiqihar,Harbin,Jiamusi,Mudanjiang.Foot dermatitis rate was highest in Mudanjiang area.Deformed hoof,ulcers on the sole and tip of the hoof,white line disease and rotting hoof disease were the highest in Jiamusi area.The incidence of laminitis is high est in Daqing area.The incidence of limb and hoof diseases in large pasture(26.9%)was lower than that in small pasture(35.0%)and medium pasture(28.7%).The seasonal factors,environmental factors,bed factors,hoof protection factors,body condition sco re,number of parities,lactation volume and lactation period all had effects on limb and hoof diseases in dairy cows.Among them,seasonal factors have the greatest influence on limb and hoof diseases,the highest incidence of limb and hoof diseases in winter(8.7%),followed by summer(7.5%),spring and autum was 6.2%?5.9%,respectively.Compared with pastures with clean ground,factors such as poor sanitation and low frequency of scraping manure will increase the incidence of limb and hoof diseases.The incidence of limb and hoof diseases in poorly hygienic cattle farms(35.9%)is significantly higher than that in well-hygienic cattle farms(23.5%),but the different types of bedding materials(Sand,rice husk,rice husk + fermented cow dung)have no significant impact on cow limb and hoof diseases.The frequency of hoof protection could also significantly affect the incidence of limb and hoof diseases.According to the survey,the incidence of limb and hoof diseases was 25.8% in the pastures that repaired feet twice a year+daily,and 31.9% in the pastures that repaired feet only twice a year.The incidence of limb and hoof diseases in the pastures that bathed once a week(35.9%)was also higher than that in the pastures that bathed feet three times(24.0%).In addition,the comprehensive comparative analysis showed that the incidence of limb and hoof diseases in cows with abnormal body weight was significantly higher than that in the normal group(28.2%),which indicated that weight management was also one of the important factors for daily preven tion of hoof disease,whether it was emasculated(32.3%)or obese(35.9%).In this survey,we found that the incidence of limb and hoof diseases increased with the increase in parity.The incidence of limb and hoof diseases in dairy cows with one child was 24.7%,and the incidence of limb and hoof diseases in five or more cows reached 34.0%.The lactation amount is also significantly proportional to the incidence of limb and hoof diseases.The incidence of limb and hoof diseases in dairy cows with a lactation <7.5 t is 26.0%,and the incidence of limb and hoof diseases is as high as 31.4% in dairy cows with a lactation yield of more than 10.5 t.In addition,in different stages of lactation,the incidence of limb and hoof diseases has changed significantly,the highest incidence of limb and hoof diseases in early lactation is 9.3%,with the decrease of lactation,the incidence of limb and hoof diseases decreases,and the incidence of limb and hoof diseases in late lactation is7.4%.Dairy cows had the lowest incidence during the perinatal period(3.0%).Screening the undetermined markers through the rat osteoarthritis model found that within 10 weeks of surgically induced rat osteoarthritis model,the pathological changes of tibial cartilage gradually increased,the cartilage surface was damaged,and the middle and deep chondrocytes were vacuolated.At the 10 th week after the operation,chondrocytes were lost and the subchondral bone was severely damaged.Detecting the contents of various related markers in rat serum within10 weeks by ELISA found that the concentrations of GAG and PIICP did not change significantly,while the concentrations of CS846,COMP,C2 C and CTX-II markers showed a significant increasing trend with the course of the disease,which was pos itively correlated with the degree of joint damage.Also,compared with healthy cows,serum CS846?COMP?C2C?CTX-II markers were significantly increased in cows with osteoarthritis(P<0.05),suggesting that these four indicators can be used as sensitive biomarkers for osteoarthritis.Compared with the contro l group,the serum activity of MMP9 was significantly increased(P<0.05),but the serum activity of TIMP1 was decreased at each time period,there was no significant difference.Suggesting that MMP9 could be used as potential biomarker and guide the early diagnosis of cow laminitis.This study identified the epidemic characteristics and rules of cow limb and hoof diseases in Heilongjiang Province,screen the sensitive characteristic markers of cow osteoarthritis and cow laminitis,and provide theoretical basis for the rapid diagnosis of cow osteoarthritis and cow laminitis.It provides scientific reference and new ideas for the overall prevention and control of cow limb and hoof diseases in Heilongjiang province,and is of great significance for improving the overall diagnosis and treatment level of cow limb and hoof diseases and the economic benefits of the industry. |
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