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1.
Bovine viral diarrhea viruses (BVDV) cause both acute and persistent infections. While diagnostic tests have been designed to detect animals persistently infected (PI) with BVDV, the reliability of these tests in detecting acute BVDV infections is not known. It is also possible that acute BVDV infections may be confused with persistent infections in surveys for PI animals. In this study, 2 tests presently in use in diagnostic laboratories to test for PI animals, polymerase chain reaction amplification followed by probe hybridization (RT-PCR/probe) of serum samples and immunohistochemical detection of viral antigen in skin biopsies (IHC), were evaluated for their ability to detect acute BVDV infections. Sixteen colostrum-deprived, BVDV-free, and BVDV-antibody-free calves were infected with 6 different BVDV strains. Clinical signs, seroconversion, and virus isolation indicated that inoculated animals did replicate virus. Virus could be detected in 19% (3/16) of acutely infected animals by the RT-PCR/probe technique. No acutely infected animals were positive by IHC.  相似文献   

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The prevalence of bovine viral diarrhea virus (BVDV) in persistently infected (PI) cattle in beef breeding herds was determined using 30 herds with 4530 calves. The samples were collected by ear notches and tested for BVDV antigens using immunohistochemistry (IHC) and antigen capture enzyme-linked immunosorbent assay (ACE). Animals with initial positives on both IHC and ACE were sampled again using both tests and serums were collected for viral propagation and sequencing of a viral genomic region, 5′-untranslated region (5′-UTR) for viral subtyping. Samples were also collected from the dams of PI calves. There were 25 PI calves from 4530 samples (0.55%) and these PI calves were from 5 of the 30 herds (16.7%). Two herds had multiple PI calves and 3 herds had only 1 PI calf. Only 1 of the 25 dams with a PI calf was also PI (4.0%). The subtype of all the PI isolates was BVDV1b. Histories of the ranches indicated 23 out of 30 had herd additions of untested breeding females. Twenty-four of the 30 herds had adult cowherd vaccinations against BVDV, primarily using killed BVDV vaccines at pregnancy examination.  相似文献   

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OBJECTIVE: To report the prevalence of bovine viral diarrhea virus (BVDV) in calves and calf groups (ie, calves from the same farm) in beef breeding herds and evaluate the ability of biosecurity risk assessment questionnaires to identify calf groups with positive results for BVDV. DESIGN: Nonrandom survey. ANIMALS: 12,030 calves born in spring from 102 operations. PROCEDURES: Cow-calf producers that voluntarily enrolled in a screening project submitted ear notch specimens from calves and answered a 29-question survey instrument. Ear notch specimens were tested for BVDV with an antigen-capture ELISA (ACE), and ear notch specimens with positive ACE results for BVDV were immediately retested by performing immunohistochemistry (IHC). Follow-up testing, 3 to 4 weeks after initial positive ACE results, was done by use of a second IHC test and virus isolation on a subsequently submitted ear notch specimen from the same calves to identify those that were persistently infected (PI). RESULTS: 102 producers submitted ear notch specimens for BVDV screening. Initially, 24 of 12,030 calves had positive ACE results for BVDV. A second ear notch specimen was submitted for 20 of these 24 calves. Of 20 retested calves, 12 had positive ICH results for BVDV, confirming PI status. The 12 PI calves came from 4 calf groups (3 singletons and 1 calf group with 9 PI calves). CONCLUSIONS AND CLINICAL RELEVANCE: Prevalence of BVDV in calf groups was low, and questions designed to identify high-risk biosecurity behaviors had little value in identifying calf groups with positive results for BVDV.  相似文献   

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Calves persistently infected (PI) with Bovine viral diarrhea virus (BVDV) represent an important source of infection for susceptible cattle. We evaluated vaccine efficacy using calves PI with noncytopathic BVDV2a for the challenge and compared tests to detect BVDV in acutely or transiently infected calves versus PI calves. Vaccination with 2 doses of modified live virus vaccine containing BVDV1a and BVDV2a protected the calves exposed to the PI calves: neither viremia nor nasal shedding occurred. An immunohistochemistry test on formalin-fixed ear notches and an antigen-capture enzyme-linked immunosorbent assay on fresh notches in phosphate-buffered saline did not detect BVDV antigen in any of the acutely or transiently infected calves, whereas both tests had positive results in all the PI calves.  相似文献   

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Research aimed at optimising diagnostic laboratory procedures is central to the development of effective bovine viral diarrhoea virus (BVDV) control programmes. BVDV is a single-stranded RNA virus that crosses the placenta to infect foetuses, resulting in reproductive losses due to foetal death or persistently infected calves that die early in life. Persistently infected animals are widely accepted to be the primary reservoir of BVDV and the largest source of infection. This poses important challenges to overall animal/herd health and can cause major losses to the cattle industry. Long-term storage of bovine ear notch samples from calves persistently infected with BVDV may adversely affect the ability of diagnostic assays to detect the virus efficiently. In order to test this hypothesis, ear notch samples from 7 animals were divided into 2 groups. One set was subjected to prompt formalin fixation and the other set stored either as fresh samples without preservatives at -2 degrees C, or soaked overnight in phosphate buffered saline followed by freezing of the supernatant fluid at -2 degrees C. Frozen ear notches and ear notch supernatant yielded positive results with an antigen-capture, enzyme linked immunosorbent assay (AC-ELISA) for the duration of the study (6 months) and optical density (OD) values remained significantly within range. There was no significant difference between storing fresh ear notch samples or PBS at -2 degrees C. However, positive immunohistochemistry (IHC) staining on formalin fixed ear notches started to fade between Day 17 and Day 29 when stored at room temperature. It was concluded that fresh ear notches could safely be stored at -2 degrees C for a period of 6 months prior to testing for BVD viral antigens.  相似文献   

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Bovine viral diarrhea virus (BVDV) persistently infected (PI) calves represent significant sources of infection to susceptible cattle. The objectives of this study were to determine if PI calves transmitted infection to vaccinated and unvaccinated calves, to determine if BVDV vaccine strains could be differentiated from the PI field strains by subtyping molecular techniques, and if there were different rates of recovery from peripheral blood leukocytes (PBL) versus serums for acutely infected calves. Calves PI with BVDV1b were placed in pens with nonvaccinated and vaccinated calves for 35 d. Peripheral blood leukocytes, serums, and nasal swabs were collected for viral isolation and serology. In addition, transmission of Bovine herpes virus 1 (BHV-1), Parainfluenza-3 virus (PI-3V), and Bovine respiratory syncytial virus (BRSV) was monitored during the 35 d observation period. Bovine viral diarrhea virus subtype 1b was transmitted to both vaccinated and nonvaccinated calves, including BVDV1b seronegative and seropositive calves, after exposure to PI calves. There was evidence of transmission by viral isolation from PBL, nasal swabs, or both, and seroconversions to BVDV1b. For the unvaccinated calves, 83.2% seroconverted to BVDV1b. The high level of transmission by PI calves is illustrated by seroconversion rates of nonvaccinated calves in individual pens: 70% to 100% seroconversion to the BVDV1b. Bovine viral diarrhea virus was isolated from 45 out of 202 calves in this study. These included BVDV1b in ranch and order buyer (OB) calves, plus BVDV strains identified as vaccinal strains that were in modified live virus (MLV) vaccines given to half the OB calves 3 d prior to the study. The BVDV1b isolates in exposed calves were detected between collection days 7 and 21 after exposure to PI calves. Bovine viral diarrhea virus was recovered more frequently from PBL than serum in acutely infected calves. Bovine viral diarrhea virus was also isolated from the lungs of 2 of 7 calves that were dying with pulmonary lesions. Two of the calves dying with pneumonic lesions in the study had been BVDV1b viremic prior to death. Bovine viral diarrhea virus 1b was isolated from both calves that received the killed or MLV vaccines. There were cytopathic (CP) strains isolated from MLV vaccinated calves during the same time frame as the BVDV1b isolations. These viruses were typed by polymerase chain reaction (PCR) and genetic sequencing, and most CP were confirmed as vaccinal origin. A BVDV2 NCP strain was found in only 1 OB calf, on multiple collections, and the calf seroconverted to BVDV2. This virus was not identical to the BVDV2 CP 296 vaccine strain. The use of subtyping is required to differentiate vaccinal strains from the field strains. This study detected 2 different vaccine strains, the BVDV1b in PI calves and infected contact calves, and a heterologous BVDV2 subtype brought in as an acutely infected calf. The MLV vaccination, with BVDV1a and BVDV2 components, administered 3 d prior to exposure to PI calves did not protect 100% against BVDV1b viremias or nasal shedding. There were other agents associated with the bovine respiratory disease signs and lesions in this study including Mannheimia haemolytica, Mycoplasma spp., PI-3V, BRSV, and BHV-1.  相似文献   

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The objective of this study was to evaluate the efficacy of immunohistochemical (IHC) staining for diagnosis of persistent bovine viral diarrhea virus (BVDV) infection using formalin-fixed, paraffin-embedded skin biopsy specimens. Skin from 41 of 42 calves shown to be persistently infected (PI) with BVDV by repeated virus isolation more than 3 weeks apart were immunohistochemically positive for BVDV antigen. Positive IHC staining was most pronounced in the keratinocytes and in hair follicle epithelium, hair matrix cells of the hair bulb, and the dermal papilla. All of the skin sections from 10 calves experimentally infected postnatally with BVDV (10(5) median tissue culture infective doses [TCID50]) and biopsied on days 0, 5, 7, and 9 postinfection were negative for viral antigen. Ten calves from a second group experimentally infected with a higher dose of BVDV (10(8) TCID50) were biopsied when viremic between 10 and 14 days postinfection and 4 calves exhibited positive IHC staining for BVDV; however, staining in these skin biopsies was confined to small foci in the nonfollicular epidermis and follicular ostia. This staining was distinct from that observed in skin obtained from PI cattle. Skin biopsy represents an effective method for identifying animals PI with BVDV.  相似文献   

13.
Viral distribution and lesions were compared between calves born with persistent infection (PI) and calves acutely infected with the same bovine viral diarrhea virus (BVDV) isolate. Two PI calves from 1 dairy herd were necropsied. The PI viruses from these calves were isolated, characterized by sequencing, and found to be identical. This virus strain, designated BVDV2-RS886, was characterized as a noncytopathic (ncp) type 2 BVDV. To establish acute infections, BVDV2-RS886 was used to inoculate clinically healthy, seronegative calves which were 3 weeks to 3 months old. Nine calves received 10(6)-10(7) tissue culture infective dose of BVDV2-RS886 intranasally. Four additional age-matched animals served as noninfected controls. Infected calves were necropsied at 3, 6, 9, or 13 days postinoculation (dpi). Viral antigen was detected by immunohistochemistry in frozen sections, and lesions were evaluated in hematoxylin eosin-stained paraplast sections. In the PI calves, a wide distribution of viral antigen was found in all tissues and was not associated with lesions. In the acutely infected calves, viral antigen was widespread in lymphoid tissues at 6 dpi but had been mostly eliminated at 9 and 13 dpi. Depletion of lymphoid tissues was seen at 6, 9, and 13 dpi and repopulation at 9 and 13 dpi. In 1 of the calves at 13 dpi, severe arteritis was present in lymph nodes and myocardium. This comparison shows that an ncp BVDV strain that causes no lesions in PI animals is able to induce marked depletion of lymphoid tissues in calves with acute infection. Therefore, the failure to eliminate PI cattle from a herd causes problems not only in pregnant cattle but may also affect other age groups.  相似文献   

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A new sandwich enzyme-linked immunosorbent assay (S-ELISA) kit that uses raw (unprocessed) fetal bovine serum (FBS) as the testing sample was evaluated for upstream bovine viral diarrhea virus (BVDV) testing. Pooled FBS samples (n = 84) were tested using the S-ELISA. Thirty serum samples originating from persistently infected (PI) calves that had been confirmed by virus isolation (VI) as BVDV positive and another 30 samples previously confirmed by VI as BVDV negative were also evaluated. Of the 84 field samples, the S-ELISA detected 13 (15.5%) BVDV-positive specimens. When these 13 positive samples were tested by VI and immunofluorescent assay, 11 (84.6%) were positive and 2 (15.4%) were negative. The S-ELISA was positive for all 30 PI samples (100%) and negative for all 30 negative samples (100%). These data indicate that the new kit is a relatively reliable diagnostic tool and can be considered for upstream detection of BVDV-contaminated raw FBS pools.  相似文献   

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In the summer of 1996, we screened 18,931 calves in 128 beef herds located in five US states for persistent bovine viral diarrhea virus (BVDV) infection. Of these, 76 herds were randomly selected from the client database of collaborating veterinary practices, and 52 herds were suspected by the collaborating veterinarians to have BVDV infection based on history or clinical signs. Serum was obtained from each calf in the cooperating herds prior to 4 months of age and tested for the presence of BVDV by microtiter virus isolation. Information about each of the herds (including management practices, vaccination history, and breeding- and calving-season production measures) were collected by the collaborating veterinarians using standardized questionnaires. A total of 56 BVDV-positive calves in 13 herds were identified on initial screening. Ten (19%) of the BVDV-suspect herds and three (4%) of the randomly selected herds had > or = 1 BVDV-positive calf at initial screening. Multiple BVDV-positive calves were identified in 10 of those 13 herds. Follow-up information was obtained for 54 of the 56 positive calves. Ten out of 54 (18%) died prior to weaning, and 1 (2%) was sold because of unusually poor growth. Thirty-three out of 54 (61%) of the initially positive calves remained BVDV positive at 6 months of age - confirming persistent-infection (PI) status. Dams of 45 of the 56 positive calves were tested, with 3 (7%) identified as positive - indicating most PI calves were products of acute dam infection during gestation. The proportion of cows that were pregnant at the fall 1995 pregnancy examination was 5% lower in herds with PI calves born during the 1996 calving season than in randomly selected herds without PI calves. Most of the calves we identified with persistent BVDV infections survived to weaning, and could provide a constant source of virus to the herd throughout the breeding season and early gestation.  相似文献   

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Bovine viral diarrhea virus (BVDV) infections resulting in clinical disease developed in calves, despite vaccination of dams and high maternal BVDV antibody titers in calves. Eight persistently infected (PI) calves born to immunocompetent dams were identified in the herd. Neutralizing BVDV antibody titers of PI calves had decreased greatly by the time the calves were 1 to 2 months old. Antibody titers of PI calves decreased more rapidly than antibody titers of calves that were not PI. Reduced antibody titers in PI calves allowed detection of BVDV in serum specimens of all PI calves by the time they were 8 weeks old. Persistent infection in suspect calves was detectable serologically and was confirmed by virologic examination of serum specimens 4 months after weaning, when the calves were 9 months old. Growth rates were reduced in viremic calves.  相似文献   

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A new diagnostic approach testing tissue samples derived from cattle ear tagging for bovine viral diarrhoea virus (BVDV) antigen in a commercially available antigen capture enzyme-linked immunosorbent assay (ACE) was developed. To validate this method, 99 positive and 469 negative samples were tested. With those samples the assay yielded a sensitivity of 100% and specificity of >or=99.6%. Serum and ear tissue samples from 11 persistently infected (PI) BVDV calves were tested. While serum samples were negative after intake of colostrum, the ear tissue samples could be detected positive for BVDV all the time. Testing multiple samples derived from the same ear from PI cattle yielded positive results and low variation. Using cattle ear tags combining the ear tag application with sampling of a small ear tissue plug and testing those tissue samples with an ACE could be a reliable and economic way of BVDV testing.  相似文献   

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Bovine viral diarrhea virus (BVDV) affects cattle populations causing clinical signs that range from subclinical immunosuppression to severe reproductive and respiratory problems. Detection and removal of persistently infected (PI) calves is the single most important factor for control and eradication of BVDV. Current testing strategies to detect PI calves rely heavily on immunohistochemistry (IHC) and a commercially available antigen capture ELISA (ACE) assay. These viral assays depend on 1 or 2 monoclonal antibodies which target the E(rns) glycoprotein of BVDV. The sensitivity and specificity of these two tests have been reported previously. The purpose of this research was to characterize a strain of BVDV (AU501) that was undetectable using IHC and ACE based on a single monoclonal antibody, but was consistently detected in samples from a Holstein steer using virus isolation and PCR testing. Sequencing of this AU501 viral isolate revealed a unique mutation in the portion of the genome coding for the E(rns) glycoprotein. This unique field strain of BVDV demonstrates the risk of relying on a single monoclonal antibody for detection of BVDV. Multiple testing strategies, including polyclonal or pooled monoclonal antibodies that detect more than one viral glycoprotein may be necessary to detect all PI calves and facilitate eradication of BVDV.  相似文献   

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