A retrospective study into characteristics associated with the seroprevalence of pseudorabies virus-infected breeding pigs in vaccinated herds in the southern Netherlands

Vaccination programs to eradicate pseudorabies virus (PRV) are being considered in several countries. Knowledge of factors that influence PRV transmission within vaccinated breeding herds may contribute to the success of these programs. A multivariate analysis of variance of the PRV-seroprevalence in sows in 209 herds (average seroprevalence 67.0% per herd) in the southern Netherlands revealed the following risk indicators: (1) presence of finishing pigs; (2) production type (producers of finishing piglets had a higher seroprevalence than producers of breeding stock); (3) vaccination of the sows during nursing (in comparison with vaccinating all sows simultaneously at 5 month intervals, or vaccination during the second half of gestation); (4) pig density in the municipality where the herd was located (seroprevalence increased with higher pig density); (5) herd size less than 100 sows; (6) average within-herd parity (seroprevalence increased with higher withinherd parity); (7) replacement pigs raised on the premises; (8) vaccine strain administered to the sows. Purchase policy (breeding pigs purchased between 10 weeks and 7 months of age, or use of home-bred gilts only) did not significantly contribute to the multivariate model.     

Evaluation of pseudorabies viral antigens in the agar gel immunodiffusion test.

Procedures designed to extract pseudorabies viral (PRV) antigens from PRV-infected tissue cultures were investigated to determine whether differences in extraction method had an effect upon the final concentrated antigenic product. All four of the preparations made from PRV-infected tissue culture cells (trypsin extract and disrupted cells) or entire PRV-infected cultures (polysorbate 80 extract and (NH4)2SO4 precipitate) contained relatively large amounts of the same antigen, whereas cell-free PRV-infected tissue culture fluids did not contain significant amounts of this antigen. Specific antibody directed against this antigen was present in all PRV antisera tested. Two other antigens were observed in some of the preparations, but PRV antisera varied in their ability to precipitate with these antigens. Therefore, the number of precipitation lines observed in agar gel immunodiffusion between PRV preparations and PRV-positive antisera depended both upon the extraction method used to obtain the antigen and upon the specificity of the selected antiserum.     

Potential role of viral surface glycoproteins in the replication of H3N2 triple reassortant influenza A viruses in swine and turkeys

The H3N2 triple reassortant (TR) influenza viruses emerged in swine in 1998 and then in turkeys in 2003. It was then hypothesized that these viruses crossed the species barrier and transmitted from pigs to turkeys. In previous work we identified viruses with different transmission behavior between the two species, of which A/turkey/Ohio/313053/04 (TK04) transmitted both ways between swine and turkeys, and A/swine/North Carolina/03 (SW03) did not transmit either way between the two species. Utilizing the 12-plasmid reverse genetics (RG) system, we rescued two viruses (TK04 and SW03) with potentially different transmission behavior between pigs and turkeys. Single gene reassortants (SGR) were generated by switching the hemagglutinin (HA) or the neuraminidase (NA) genes between both viruses, and were evaluated for replication in vitro (pig and turkey tracheal/bronchial epithelial cells) and in vivo (pigs and turkeys). RG-created TK04 replicated more efficiently than SW03 in vitro and in vivo. Additionally, TK04 exhibited better binding affinity to plasma membrane preparations (PMP) from pig and turkey tracheal/bronchial epithelial cells compared to SW03. In study with SGR viruses, the HA protein was found to be essential for TK04 virus transmission amongst turkeys, but not sole factor contributing to the efficient replication of virus in turkeys and pigs. Such findings further highlight the polygenic nature of influenza virus pathogenesis.     

Detection of viral antigens in bluetongue virus-infected ovine tissues, using the peroxidase-antiperoxidase technique

An indirect immunoperoxidase procedure was developed to detect viral antigens in bluetongue virus (BTV)-infected tissues. Embryonating chicken eggs were infected with BTV serotypes 10, 11, 13, or 17, and the chorioallantoic membranes were subsequently fixed in formalin and embedded in paraffin. The peroxidase-antiperoxidase (PAP) system was used to examine the infected membranes for the presence of viral antigens. Sheep antisera raised against BTV serotypes 10, 11, 13, and 17 served as the primary antibodies in the PAP procedure. Specific staining was observed when each of these antisera was applied to membranes expressing antigens of homologous and heterologous BTV serotypes. The PAP method was rapid, reliable, and specific in its detection of BTV.     

The fetal brain in bovine viral diarrhea virus-infected calves: lesions, distribution, and cellular heterogeneity of viral antigen at 190 days gestation

Previous studies have shown that the brain is a target of persistent infection with bovine viral diarrhea virus (BVDV) and have demonstrated viral tropism for neurons as well as other endogenous cell types in diverse brain areas. Apart from foci of mild residual inflammation in some postnatal calves, consistent brain lesions, per se, have not been reported. No similar comprehensive studies of the brain have been reported in bovine fetuses. In the current study, 12 BVDV-seronegative heifers were inoculated intranasally with a 2-ml 4.4 log(10) TCID(50)/ml dose of noncytopathic type 2 BVDV at 75 and 175 days of gestation to create persistently and transiently infected fetuses, respectively. In only persistently infected fetuses, encephaloclastic lesions resulting in pseudocysts were observed in the subependymal zone in the region of the median eminence and adjacent corona radiata as well as in the region of the external capsule associated with lenticulostriate arteries. Additionally, areas of rarefaction in white matter were observed at the tips of cerebrocortical gyri and in the external capsule. The distribution of viral antigen was examined by immunohistochemical labeling using the 15C5 anti-BVDV monoclonal antibody. Viral antigen was detected only in calves inoculated at 75 days of gestation, i.e., persistently infected. The pattern of BVDV immunolabeling revealed both similarities and differences compared with previous studies in postnatal calves, suggesting that viral infection in the brain is a dynamic and progressive rather than static process.     

Role of the raccoon in the transmission of pseudorabies: a field and laboratory investigation

Serum samples were collected from 42 raccoons trapped in a pseudorabies enzootic area of Missouri. All samples were negative for neutralizing antibodies to pseudorabies virus (PRV). Raccoons were orally exposed with one of four dose levels of PRV. The raccoons were found to be susceptible to moderately large doses of PRV tissue culture infective dose (10(4) - 10(5) TCID50). All raccoons given 10(3) TCID50 of PRV survived and did not develop clinical signs. The PRV was consistently isolated from tonsillar swabs collected from raccoons that died after exposure. The PRV was also isolated from the brain, tonsils, lungs, and salivary glands of raccoons that died. It was isolated from samples of the brain and tonsils collected from a raccoon 5 days after death. Neutralizing antibodies to PRV could not be detected in the serum of raccoons that survived PRV exposure. Tonsillar swabs and tissue samples collected from these raccoons were free of PRV. The results indicated that the raccoon may serve as a short-term reservoir for PRV, but it is unlikely to have an ipizootiologic role as a long-term, subclinical carrier of the virus.     

Immunogenic properties of a bovine herpesvirus-1 (BHV-1) recombinant expressing major pseudorabies virus (PrV) glycoproteins in combination

A recombinant bovine herpesvirus type 1 (BHV-1), designated BHV-1/TF17-1, which expresses pseudorabies virus (PrV) glycoproteins gB, gC, gD, gE and gI in combination was constructed. To test the protective immunity, 10 mice were inoculated with BHV-1/TF17-1 and three weeks later 10 mice were intraperitoneally (i.p.) challenged with 20 LD50 virulent PrV (YS-81). BHV-1/TF17-1 protected all the mice from the PrV lethal challenge while all the control mice died in around 3 days. Mice vaccinated with BHV-1/TF17-1 acquired high PrV-neutralizing antibody titers and demonstrated strong delayed type hypersensitivity responses and moderate in vitro lymphocyte proliferative responses to PrV antigen. Since the major PrV glycoproteins were integrated into virions (probably into viral envelope), BHV-1/17-1 was neutralized with anti-PrV antiserum. However, the susceptibility of BHV-1/TF17-1 to anti-PrV antiserum is 2- to 4-fold lower than that of PrV vaccine lines. Our results demonstrated the possibility of BHV-1/17-1 as a vaccine to protect piglets from Audjesky's disease where maternal antibodies against PrV interfere attenuated live PrV vaccines.     

A study comparing the immunologic responses of swine to pseudorabies viral antigens based on the ELISA, serum virus neutralization, and latex agglutination tests

A study of pseudorabies virus (PRV)-vaccinated pigs comparing the immune responses detected by the latex agglutination test (LAT) with responses detected by other routine tests for pseudorabies antibodies indicated that LAT was more sensitive than either the enzyme-linked immunosorbent assay (ELISA) or the serum virus neutralization test (SVNT). The LAT detected antibodies sooner than ELISA and SVNT in unvaccinated pigs after challenge with virulent PRV. The specificities of the 3 tests were found to be near 100%. The LAT is a good alternative to SVNT or ELISA for detection of PRV-specific antibodies.     

Demonstration of pseudorabies viral antigen in thick and ultrathin tissue sections of young pigs using an immunogold method

Pseudorabies (Aujeszky's) virus antigens were labeled in thick and ultrathin tissue sections of young pig brain and liver tissue, using an indirect immunogold method. Antigens were tagged with 20 nm gold particles. Controls proved the specificity of the reaction in paraffin sections and ultrathin epoxy sections. Immunogold staining was compared with immunoperoxidase staining in paraffin sections. In ultrathin sections stained with the immunogold method, the gold particles were present on viral nucleocapsids and viral envelopes, as well as on a number of other intracellular structures. These included the inner nuclear membrane, the nucleoplasm, intranuclear filaments, the endoplasmic reticulum, and free cytoplasmic polyribosomes. Gold particles were absent on mitochondria and microtubules. In paraffin sections, immunogold labeling for pseudorabies virus antigen was less sensitive than immunoperoxidase staining. Immunogold staining of ultrathin tissue sections can yield additional information on virus-host cell interactions.     

猪伪狂犬病病毒疫苗株Bartha-K61在犬科动物体内的安全性评价及免疫实验

为了对猪伪狂犬病病毒(PRV)疫苗株Bartha-K61在犬科动物体内的安全性和免疫原性作出评价,通过3种免疫途径(肌肉注射、滴鼻免疫和口服免疫)对犬进行免疫(2 mL,6×10 8PFU/mL),并在免疫结束后通过组织病理切片、免疫组化、临床表现、产仔性能、排泄物中病毒存留情况进行了检测.结果发现,在PRV易感器官内未发现病毒抗原,且疫苗接种犬体温正常,精神状态良好,产仔正常,无任何不良症状,排泄物中亦无病毒粒子残留;同时,通过对免疫后犬只的细胞免疫水平、体液免疫水平和中和试验的检测发现,3种免疫途径均可有效地刺激机体产生免疫应答,且中和抗体水平达到较高水平.从而证明,PRV Bartha-K61在犬科动物体内安全有效,为以PRV为栽体表达犬科动物疫病的抗原基因重组活栽体疫苗的研制提供了科学依据.     

表达猪细小病毒VP2基因的重组伪狂犬病病毒构建及其免疫原性

猪细小病毒(porcine parvovirus,PPV)和伪狂犬病病毒(pseudorabies virus,PRV)均可引起母猪的繁殖障碍疾病。本试验以PRV基因工程弱毒株rPRVSMX为载体,构建了表达PPV流行毒株VP2基因的重组PRV(rPRVSMX-VP2)。Western blot和IFA试验均证明了重组病毒在细胞中成功表达了VP2蛋白。动物试验结果证明重组病毒可以诱导猪体产生特异性的PPV抗体,首免后40 d血凝抑制抗体(HI)为1∶192±73.9,虽然低于PPV商品化灭活疫苗免疫后所产生的HI抗体,但此时免疫系统被认为是激活的水平;重组病毒诱导产生的PRV抗体与载体毒株rPRVSMX诱导产生的抗体相当。由此可见该重组毒株经过优化后,可以作为二联疫苗的候选毒株防控猪伪狂犬病和细小病毒病。     

The role of TREM-2 in internalization and intracellular survival of Brucella abortus in murine macrophages

Triggering receptor expressed on myeloid cells-2 (TREM-2) is a cell surface receptor primarily expressed on macrophages and dendritic cells. TREM-2 functions as a phagocytic receptor for bacteria as well as an inhibitor of Toll like receptors (TLR) induced inflammatory cytokines. However, the role of TREM-2 in Brucella intracellular growth remains unknown. To investigate whether TREM-2 is involved in Brucella intracellular survival, we chose bone marrow derived macrophages (BMDMs), in which TREM-2 is stably expressed, as cell model. Colony formation Units (CFUs) assay suggests that TREM-2 is involved in the internalization of Brucella abortus (B. abortus) by macrophages, while silencing of TREM-2 decreases intracellular survival of B. abortus. To further study the underlying mechanisms of TREM-2-mediated bacterial intracellular survival, we examined the activation of B. abortus-infected macrophages through determining the kinetics of activation of the three MAPKs, including ERK, JNK and p38, and measuring TNFα production in response to lipopolysaccharide (LPS) of Brucella (BrLPS) or B. abortus stimulation. Our data show that TREM-2 deficiency promotes activation of Brucella-infected macrophages. Moreover, our data also demonstrate that macrophage activation promotes killing of Brucella by enhancing nitric oxygen (NO), but not reactive oxygen species (ROS) production, macrophage apoptosis or cellular death. Taken together, these findings provide a novel interpretation of Brucella intracellular growth through inhibition of NO production produced by TREM-2-mediated activated macrophages.     

某规模化猪场伪狂犬病抗体检测与分析

目的:为了掌握规模化猪场伪狂犬病野毒感染情况,减少伪狂犬病所造成的经济损失,以及规模化猪场应当采取的有效防疫措施。方法:采用ELISA法对某规模化猪场7~12月份的经产母猪和后备猪进行伪狂犬病抗体检测。结果:该猪场的伪狂犬病阳性率为21.26%,是伪狂犬病野毒阳性场。结论:该猪场在检测出伪狂犬病抗体呈阳性后,采取了一系列措施,从而控制了疫情的发生。     

沈阳某猪场伪狂犬病流行病学调查及免疫效果评估

采用猪伪狂犬病毒抗体检测试剂盒,对沈阳某使用gE基因缺失疫苗免疫的生猪养殖场进行猪伪狂犬病毒抗体检测,同时使用PCR方法对猪伪狂犬病毒gE基因进行筛查,对该场的疫苗免疫效果及病毒隐性感染情况进行评估。结果表明,该场猪伪狂犬病毒隐性感染率为11.5%;疫苗免疫效果母猪最佳,哺乳仔猪次之,育肥猪再次。建议该场根据试验结果调整免疫程序,并对隐性感染病猪及时进行淘汰、净化。