| Infectious bovine rhinotracheitis?IBR?,also called "red nose",is an acute,febrile and contagious disease of cow caused by infectious bovine rhinotracheitis virus?IBRV?.There is often secondary onset of manifold bacterial infections to the cattle 5-7d later after infected with acute IBR,especially the newborn calves may suffer from secondary onset of infection of manifold bacterium such as colibacillus,Pasteurella,and streptococci,etc.leading to high incidence and mortality of calves.In recent years,some scalized cow ranch in Xinjiang,many calves suffered from suspected infectious bovine rhinotracheitis followed by secondary onset of bacterial meningitis,causing large economic losses to the cow ranch.For this,the research firstly made an epidemiological survey in the field to find out the possible aetiological agent and potential infection factors,etc.,then acquired specimen of diseased calves in an aseptic way to detect nucleic acid of IBRV via PCR method and made a sequence analysis;what follows was separating,authenticating and making drug sensitive test for secondary infectious bacteria by combining conventional microbiology with molecular biology,and observing the experiment of isolates infecting mice;lastly a pertinent comprehensive scheme of control and prevention was established according to result of field epidemiological survey and detection,including immunizing test of comparison for vaccine,etc.The test result is as follows:1.Field epidemiological surveyThe result of field epidemiological survey showed that this disease occurs and infects quickly,and has two modes of vertical transmission and horizontal transmission,with a delitescence of 3-10 days,incidence of up to 39.64%,mortality of up to 64.18%,cure rate of 26.87% and mortality rate of 8.96%.Wherein the calves aged 0 day and 1-30 days accounted for 10.65% and 15.98% in incidence,the calves aged between 31-120 days accounted for 13.01% in total incidence;the mortalities of calves aged 0,1-30,31-60 days were respectively 72.22%,77.78% and 57.14%.The suffering cow herd this time were mainly calves aged below 4 months,with symptoms dominated by those of respiratory tract accompanied with neurological symptoms.The necropsy found hemorrhage in such tissues as brain and lung of the died calves,and lymphadenectasis and hemorrhage in each organ.2.PCR detection of infectious bovine rhinotracheitis virus and analysis on gD gene sequencePCR method was adopted to detect gD gene of infectious bovine rhinotracheitis virus,and 9 PCR positive products were selected to make a comparative analysis on sequence for comparison of homology.Result displays the IBRV-PCR positive rates of diseased calves in three different ranches were up to 70%,positive rate for nasal cavity swab of healthy calves was up to 50%,and the positive rate for nasal cavity swab for adult cows was 35%.The IBRV homology for ranches of different regions was different,wherein the homology of 9 specimens was 74.9%-99.3%,and homology with NZLKPI01000049.1 of standard strain was 82.8%-98.2%.All detecting viruses and reference viruses were on a large evolutionary branch and had the same origin,yet certain genovariation existed between detected viruses.3.Separation,authentication,pathogenicity and drug sensitive test of enterococcus faeciumConventional microbiology method was used to separate suspected bacterium from diseased specimens of dying calves,then the morphological characteristics,cultural characteristics,hemolysis,pathogenicity,biochemical properties,molecular biology?tuf gene detection,16 S rRNA sequence comparison analysis?and medicine sensibility were detected.The result showed that the 7 isolates were all enterococcus faecium,and all amplified tuf gene fragment of Eenterococcus bacterium.Besides,the 16 S rRNA gene sequence of 7 isolates and E.faecai?mNR042054.1 entered in Genbank had homology of between 92.9%-96.5%,The homology between detecting specimens was 93%-99.2%.After combining various biological characteristics of isolates,the bacteria for them was defined as enterococcus faecium.The result of hemolysis,pathogenicity of mice and drug sensitive tests showed that on 5% sheep blood agar,4 strains showed ? hemolysis,2 strains showed ? hemolysis,and 1 strain showed no hemolysis;After intraperitoneal injection of isolates to mice,they all died within 16-26 h.The isolates were more sensitive to streptomycin,vancomycin,azithromycin,CPFX,enrofloxacin,cefotaxime sodium for injection and sulfonamide,and had certain drug resistance to tetracycline,kanamycin and gentamycin,etc.4.Contrast test of immunizing potency of vaccineThe test selected 40 adult cows aged 2 prenatal months?dry period?and broke them into two groups.One group was inoculated with BVD-MD/IBR vaccine,and another group was injected with cattle IBR vaccine and received booster immunization once after 14 days.Their blood was drawn from vena caudalis before first immunization and 20 days after immunization.The blood of calves in the tested cow herd was drawn from jugular 7 days after birth to separate serums and detect IBR antibody.The result showed that the two tests all saw infection of positive cows with wild-type virus before immunization,with positive rate being respectively 20% and 25%.In the test group of pregnant adult cows,the antibody positive rate after vaccine was inactivated by bigeminy treatment of immune BVDV/IBR was 85%,and the antibody positive rate after vaccine is inactivateed by immune IBR was 80%.The positive rates of the two groups of different vaccines for active immunity antibody of calves were respectively 75% and 63.64%.The findings showed that the pathogeny causing this onset of disease to calves was hybrid infection of infectious rhinotracheitis virus with secondary onset of enterococcus faecium A comprehensive scheme of prevention and control integrating environmental improvement,pharmacotherapy and immunization was established by combining results of field epidemiological survey,drug sensitive test and vaccine comparison to effectively control epidemics of the disease in this region. |
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