| Avian leukosis(AL)is a kind of avian tumor disease caused by avian leukosis virus(ALV)in the family Retroviridae.It is one of the three major diseases that commonly exist and induce tumors in chicken.ALV can be divided into 11 subgroups,from A to among which six subgroups,including A,B,C,D,E and J,can naturally infect chickens.The K subgroup is a newly identified ALV subgroup that has long existed in native chickens in East Asia.ALV can cause benign or malignant tumors and subclinical infection in chickens,resulting in growth retardation,egg drop and immunosuppression.Since the discovery of avian leukosis,the disease has shown a worldwide epidemic trend,resulting in huge economic losses.However,up to now,there are no effective drugs or vaccines to prevent and control AL.The most effective way to control avian leukosis is to purify the existing chicken flocks.International breeding companies have basically achieved the purification of classical exogenous ALV in the late 1980s,and the purification of ALV-J subgroup in 2005 or so.Also,a number of self-breeding egg-type chicken farms and local breed chicken farms in China have carried out the AL purification,and achieved basically the purification of exogenous ALV.However,in most yellow plumage broilers and local breeds,AL is still prevalent and spreading,which poses a great threat to the protection,development and utilization of poultry germplasm resources in China.In this study,the epidemiological situation of AL in four original chicken farms in Shanghai was investigated,and the infection dynamics and purification basis of avian leucosis were comprehensively mastered.The classical methods of ALV purification were explored,optimized and integrated,and the current commonly used detection methods and samples were improved.In combination with the specific situation of each chicken farms,the detection was continuously optimized,and with the supporting application of small group incubation and small group feeding production mode,and improvement of safety production management measures,we got a scientific,highly compatible and replicable ALV purification scheme,and carried out the purification of AL in four original chicken farms in shanghais.1.Comparison of different kits and samples for detection of ALAs a major screening method,the p27 antigen ELISA kit has been widely used in ALV purification in various countries,but it lacks the first-hand data of kit comparison.In ALV purification,different samples such as plasma,semen,egg white and cloaca cotton swab play different roles,but an evaluation of the effectiveness of each sample is lacking.In this study,we compared the positive rates of p27 antigen using three kits from domestic or foreign enterprises with different test samples.The results showed that the positive rates of egg white of brand A,B and C all were 2.08%;the positive rates of plasma cell culture medium were 10.42%,13.54%and 11.46%,respectively;and the positive rates of meconium detection were 3.13%,4.17%and 2.08%,respectively.Considering the cost and sensitivity,it is suggested that brand A or brand C kits should be selected when ELISA detecting.Cloacal cotton swab has a high false-positive rate which is not suitable for purification monitoring.There are some differences in the detection data of plasma,semen and egg white,which can complement with each other in purification monitoring.It is suggested to use the detection of p27 in meconium and egg white and DF-1 cell isolation virus in plasma and semen as the detection method for purification.This study provides a scientific basis for selection of detection methods and samples for ALV purification.2.Epidemiological investigation of avaian leukosis in four original chicken farms of Shanghai and establishment of key risk points of ALVDuring 2015-2018,the pathogen monitoring of AL in the original chicken farms of Shanghai was carried out.The results showed that the overall positive rates of p27 antigen in serum samples from four original chicken farms were 27.0%in A and B subgroups and 2.5%in J subgroup,respectively;the overall positive rate of p27 antigen in egg white samples was 0.9%and the suspicious rate was 2.3%;the positive rates in plasma and semen samples varied from 0 to 1.0%in hen plasma,and 0 to 4.0%in cock plasma.Only one farm was positive in cockerel semen,with a positive rate of 6.0%.It suggested that there existed a low level of infection of ALV in chickens.In addition,the endogenous and exogenous analysis of ALV in the original pheasant farm showed that the virus strain could not normally proliferate and passaged in DF-1 cells.It can be inferred that the pheasant population carried endogenous ALV.Sequencing analysis of the env gene showed that it had higher homology with the viral strains from the subgroups of A,J and F subgroups in GenBank.Based on endogenous property,the viral strain was determined to belong to F subgroup.Epidemiological investigation was carried out from five aspects,including basic situation,facilities and equipment,biosafety,production performance and clinical manifestation,and epidemic disease purification.It was found that the original chicken farms in Shanghai have purification foundation,and therefore the comprehensive purification of the disease should be carried forward.On the basis of full epidemiological investigation and laboratory monitoring,four key risk points for preventing the spread of AL in the original chicken farms of Shanghai were established,and corresponding control measures were formulated.This study lays a foundation for the comprehensive purification of ALV.3 Establishment of purification program for avian leukosis in Shanghai original chicken farmsIn order to control and purify AL from the source,we took an original breed chicken farm as the purification object,with the improvement of infrastructure construction(13 control points in 3 aspects),biosafety measures(21 control points in 3 aspects),production management(14 control points in 2 aspects),purification team construction(9 control points)and a good health status of poultry(2 control points)to carry out the purification of ALV.The key detection time points of purification,such as hatching day(1 day old),the late stage of brooding,the early stage of sexual maturity(caged),the early stage of laying and the preseed retention,were selected for performing the isolation of ALV in plasma and semen and detection of p27 antigen in meconium and egg white.The infected chickens were eliminated in order to block the vertical transmission of ALV,in combination with some measures to reduce horizontal transmission,such as paper bag hatching and barrier blocking in brooding stage.In the process of purification,some factors such as conservation and convenience should be taken into account.Based on the feeding mode and breeding scale of populations in the original chicken farms of Shanghai,a "one farm,one policy" purification plan was formed.4 Implementation and effect evaluation of avian leukosis purification program in four original chicken farms in ShanghaiAccording to the "one farm,one policy" purification program,the purification of AL in Shanghai original chicken farms was carried out,and a good purification result was achieved.Farm 1:After two generations of purification monitoring,the positive rates of p27 antigen in plasma of three strains at sexual maturity stage(18-20 weeks)decreased significantly from 2.0%,3.0%and 5.0%to 0.1%,0.0%and 0.1%,respectively,and the positive rates of p27 antigen in egg white and semen decreased to zero.Farm 2:After four generations of purification monitoring,the positive rate of p27 antigen in plasma decreased from 9.1%to 0.5%.And the production performance improved significantly.The average weight of 18-week-old in 7th generation increased 34.34 g in male and 19.18 g in female,the average number of eggs by 56-week-old increased 8.95,the hatching rate of hatching eggs and fertilized eggs were increased by 0.22%and 0.27%,respectively,and the survival rates of brooding,growing and laying periods were increased by 2.41%,2.17%and 0.78%,respectively.Farm 3:After one generations of purification monitoring,from seed reservation,late brooding to early laying period,the positive rates of p27 antigen in plasma varied from 2.02%,0.13%to 0.00%,the suspicious rates varied from 28.28%,0.65%to 0.24%.And the positive rate of semen remained at 0.00%,and the suspicious rate decreased from 2.13%to 0.Farm 4:A preliminary detection of ALV antigen was carried out,and the result showed that the suspicious rate was high.Due to the little number of cocks in conservation population,the suspected individuals of cocks were not eliminated in the purification process with isolating and raising separately.In conclusion,this study carried out the purification of AL in four original chicken farms in Shanghai on the basis of optimizing the detection system and integrating the purification regimen,which provides a good exemplary role in the AL purification of local and original breed chicken farms. |
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