Immunohistochemical detection of porcine teschovirus antigen in the formalin-fixed paraffin-embedded specimens from pigs experimentally infected with porcine teschovirus

Porcine teschovirus (PTV) antigens were detected by a streptavidin-biotin complex method in formalin-fixed paraffin-embedded tissues of 3-week-old pigs that had been inoculated intravenously with PTV Talfan strain. PTV antigens were detected in cytoplasm of nerve cells, glial cells and endothelial cells in the cerebellar nuclei, the grey matter of the midbrain, pons and medulla oblongata and the ventral horn of the spinal cord and of ganglion cells in the spinal ganglion corresponding to those lesions characterized as non-suppurative encephalomyelitis and ganglionitis. The results of this study suggest that nerve cells of the brain stem and spinal cord and ganglion cells of the spinal ganglion permit PTV replication and represent the main target cell population of PTV. This is the first study to demonstrate PTV antigen by immunohistochemistry in formalin-fixed paraffin-embedded tissue specimens from pigs infected with PTV.     

Optimized management of genetic variability in selected pig populations

Controlling the increase of coancestry and inbreeding coefficients in selected populations is made possible through calculation of the optimal contributions allowed to breeding animals, given the current situation with regard to genetic diversity, and further, through optimal design of matings. The potential of such an approach for pig breeding was tested by retrospective optimization on the French Landrace population in reference to the matings actually carried out during a 21-week test period. The major constraint was that the average overall estimated breeding value (EBV) should be the same as the observed one, for not decreasing short-term genetic gain. Optimizing breeding allocations to boars would have led one to decrease coancestry and inbreeding coefficients by approximately 20%. This decrease would have even increased to approximately 30%, would have replacements and disposals been optimized after accounting for genetic variability, keeping the same constraint of genetic level identical to the observed one. These results showed the potential value, in the future, of completing each periodical calculation of EBVs by optimizations considering genetic variability and of releasing corresponding information to breeders, in order to enhance maintenance of genetic variability.     

一株猪捷申病毒全基因组序列测定及分析

根据Gen Bank已发表的猪捷申病毒(porcine teschovirus,PTV)13种血清型全基因组序列,分析其保守区域并设计10对特异性引物。从临床腹泻病料中提取核酸,经RT-PCR扩增目的片段,分别对这些片段进行克隆、序列测定、拼接,最终获得一株猪捷申病毒全基因组序列,命名为PT-GD。主要结构蛋白VP1进化分析表明,该毒株与PTV12型CC25、YZ119的VP1核苷酸序列同源性分别为80.1%和79.7%,与其他血清型毒株进化距离相对较远,表明该毒株为猪捷申病毒血清型12型。该病毒全基因序列的测定及血清型鉴定为进一步深入研究猪捷申病毒在我国的遗传变异及其生物学特性提供了依据。     

Phenotypic and functional characterization of pig lymphocyte populations

Monoclonal antibodies reactive with swine lymphoid cell subpopulation antigens and with the swine major histocompatibility complex, the SLA complex, antigens are summarized. The mAb have been used to analyze swine peripheral T cell populations, the CD4+ helper T cells and the CD8+ cytotoxic/suppressor T cells. Three unique properties of swine T cells have been described. 1) There is an unusually high (25%) number of CD4+ CD8+ dual expressor T cells. 2) The ratio of CD4+ to CD8+ T cells is lower than expected; the ratio of 0.6, which is normal for pigs, only occurs in pathological conditions in humans. 3) Resting CD8+ cells preferentially express class II SLA antigens. The significance of these unusual properties of swine T cells is currently under study.     

Immunohistochemical distribution of viral antigens in pigs naturally infected with porcine teschovirus

A distribution of porcine teschovirus (PTV) antigens in pigs naturally infected with PTV is presented using the method of immunohistochemical examination. In the nervous system, PTV antigens were found in the cytoplasm of neuronal cells and glial cells distributed in the spinal ventral horn and brain stem, and also in the cytoplasm of ganglion cells in the spinal ganglion. No antigens were seen in the cerebral hemisphere. In the nervous system, the distribution of PTV antigens was consistent with lesions characteristic of nonsuppurative encephalomyelitis. In the other examined organ, PTV antigens were observed in bronchiolar epithelial cells in the lung, hepatocytes in the liver, epithelial cells in the tonsils and the myenteric nerve plexus in the small and large intestine.     

猪捷申病毒TaqMan实时定量RT-PCR方法的建立

为建立猪捷申病毒(PTV)的早期检测及定量分析方法,本研究基于PTV 11个血清型基因组5′端非编码区保守序列,设计引物和TaqMan探针,建立了检测PTV的TaqMan实时定量RT-PCR方法.应用该方法对PTV、猪细小病毒、猪繁殖与呼吸综合症病毒、猪圆环病毒2型、猪伪狂犬病毒以及猪瘟病毒进行特异性试验,结果除PTV为阳性外其它均为阴性;针对PTV最低可检测到10个拷贝;批内、批间重复试验的变异系数均小于3％.应用建立的方法与病毒分离方法分别对91份临床样品进行检测,检出率分别为79.12％和57.14％,两者的符合率是78.02％.经临床应用表明,该实时定量RT-PCR方法可为PTV的早期诊断及定量分析提供技术手段.     

猪捷申病毒Swine/CH/IMH/03株免疫原性的研究

为确定猪捷申病毒(Porcine Teschovirus,PTV)Swine/CH/IMH/03株免疫原性,本研究将该PTV分离株灭活后分别与氢氧化铝佐剂和Montanide ISA 50V2佐剂混合,制成油佐剂疫苗,并采用不同免疫程序接种实验猪后对病毒抗体进行检测。抗体检测结果表明:PTV Swine/CH/IMH/03分离株能够诱导实验动物产生较高抗体水平,并显著高于对照组;与氢氧化铝佐剂混合制成的佐剂疫苗免疫效果优于Montanide ISA 50V2佐剂;而且一次免疫组与二次免疫组没有明显差异。通过PTV Swine/CH/IMH/03株免疫原性的研究,为后续猪捷申病的预防工作提供技术支持。     

Phenotypic and genotypic traits of Staphylococcus aureus strains isolated from pig carcasses

A total of 142 S. aureus strains isolated from pig carcasses from abattoirs A (n = 98) and B (n = 44) were characterized by phenotypic and genotypic traits. Phenotypically, 96% showed yellow-pigmented colonies, 63% β/δ hemolysis, 85% were egg yolk-positive and 99% were positive for clumping factor/protein A. Only 25% of the strains were resistant to the antimicrobials tested (abattoir A: 19%; abattoir B: 39%), especially to penicillin and ampicillin. None of the strains harbored the mecA gene, which is conserved in methicillin-resistant S. aureus. The biofilm associated genes icaA, icaD and bap were present in 100%, 100% and 0% of the strains. Genes for staphylococcal enterotoxin (SE) were detected in 51% (abattoir A) and 14% of the strains (abattoir B). Among strains harboring SE genes (n = 56), 63%, 31%, 4% and 2% tested positive for seg/sei, seg, sei and sec, respectively. The amplification of the 3′ end of the coagulase gene (coa) yielded amplicons of 400, 436, 602, 682 or 776 bp. Coa restriction profile (CRP) analysis using HaeIII resulted in seven patterns (a–d, e1–e3). CRP (c) was detected most frequently at both abattoirs, whereas CRP (a) was restricted to abattoir A and CRP (e3) to abattoir B. In the slaughter process (abattoir B), (i) two CRPs (b and d) were only found before dehairing/singeing, and (ii) four CRPs (c, e1–e3) were identified throughout the process. The genotyping revealed a remarkable homogeneity in S. aureus strains from the two different abattoirs and the slaughter process stages. These results may be explained by the distribution of a limited number of S. aureus genotypes in the pig population. Moreover, as the predominant CRPs (c, e1–e3) persisted throughout the slaughter process in abattoir B, it may be hypothesized that these types are characterized by colonization advantages.     

检测猪捷申病毒抗体的间接免疫荧光方法的建立与初步应用

为建立检测猪捷申病毒(PTV)抗体的方法,本研究利用PTV-8 Jilin/2003株为杭原,建立了检All PTV杭体的的间接免疫荧光技术(IFA)方法.该方法工作浓度分别是:一抗最适稀释度为1:10:FITC标记二抗最适稀释度为1:100;二杭中伊文氏兰最适稀释度为1:30 000.敏感性试验、特异性试验表明该杭原板可以检出包含PTV-8型和1型在内的的所有PTV阳性血清,方法敏感、特异.与SN符合率94.0.用该方法对黑龙江、山东、天津和河南等地的1 500份临床血清样品进行检测,阳性检出率为55.8,表明我国猪群中PTV的感染已经非常普遍.该方法做为PTV的血清学诊断新技术,为开展PTV分子流行病学调查莫定了基础.     

Identification and molecular characterization of border disease virus (BDV) from sheep in India

Pestiviruses isolated from sheep and goats in India thus far have been bovine viral diarrhoea virus 1 (BVDV-1) or BVDV-2. During routine genetic typing of pestiviruses in the years 2009-10, border disease virus (BDV) was detected in eight Indian sheep of a flock showing clinical signs of BD by real time RT-PCR. All the samples yielded positive virus isolates in cell culture but were found negative by a BVDV antigen ELISA. A representative BDV isolate was characterized at genetic and antigenic level. Phylogenetic analysis carried out in 5′-UTR, N^pro and E2 regions of genome typed the Indian BDV isolate as BDV-3. A more detailed analysis in N^pro and entire region coding structural proteins showed that the N^pro (168), C (100 aa), E^rns (227 aa), E1 (195 aa) and E2 (373 aa) proteins were of size characteristic for BDV reference strain X818. Antigenic differences were evident between the BDV-3 isolate and previously reported BDV-1, BDV-5 and BDV-7 strains. Although origin of BDV-3 in India is not clear, the results reflect probable introduction through trade in sheep between India and other countries or BDV-3 may be more widely distributed. Additionally, this study suggests that for diagnosis of BDV infection, the commercial BVDV Ag-ELISA should be used with caution. This is the first identification of BDV in sheep in India which highlights the need for continued pestivirus surveillance and assessing its impact on sheep and goat production.     

A piglet with concurrent polioencephalomyelitis due to porcine teschovirus and postweaning multisystemic wasting syndrome

A piglet developed respiratory distress followed by difficulty in standing and unsteady gait. The lesions were characterized by polioencephalomyelitis with the predominant distribution in the brain stem, as well as lymphocyte depletion and histiocyte infiltration with cytoplasmic inclusion bodies in the lymphoid tissues throughout the body and interstitial pneumonia. Porcine teschovirus (PTV) antigens were found in the former lesions and porcine circovirus 2 (PCV2) in the latter two lesions. PTV genes were detected from the diencephalon. The results suggest that the piglet was concurrently affected with polioencephalomyelitis due to PTV and postweaning multisystemic wasting syndrome (PMWS) associated with PCV2. They also suggest that the immunosuppressive condition developing in PMWS may have facilitated the infection of the brain with PTV.     

The phenotypic and genotypic characterization of Bacillus anthracis isolates from Iran

To understand epidemiology of Bacillus anthracis in Iran, the morphological, biochemical, and virulence specifications of 32 B. anthracis isolates, collected from human, sheep, cattle, goat, and environmental specimens obtained from throughout Iran were examined by conventional and molecular approaches. B. anthracis isolates were characterized in multiple ways: (1) capsule formation both on bicarbonate agar and in defibrinated horse blood, (2) motility of vegetative forms, (3) hemolysis on 5% sheep blood agar, (4) penicillin G susceptibility, (5) lecithinase production on egg yolk agar, (6) gelatin hydrolysis, (7) ability to develop “string of pearls” on tryptose agar, and (8) capability to develop mucoid colonies in presence of CO₂ were assessed. In addition, biochemical properties such as indole, methyl red, catalase, citrate utilization, and finally nitrate reduction tests were used. All the tested isolates produced identical morphological and biochemical patterns with those of the vaccine strain B. anthracis 34F2 Sterne. In order to assess potential virulence of isolates at genomic level, PCR protocols assaying for the pXO1 and pXO2 loci were employed. The intriguing high level of phenotypic similarity between Iranian isolates of B. anthracis and the 34F2 Sterne strain deserves further studies at genomic level.     

猪捷申病毒Swine/CH/IMH/03分离株细胞适应性和致病性研究

为确定猪捷申病毒(PTV)Swine/CH/IMH/03株的易感细胞系及其致病性,本实验将该病毒接种不同细胞,通过细胞病变、电子显微镜观察和PCR技术进行鉴定。并进一步将该病毒回归本动物鉴定其致病性。试验结果表明,PTV Swine/CH/IMH/03对IB-RS-2和PK-15细胞系敏感,能够在IB-RS-2和PK-15细胞系中增殖。致病性试验结果显示,Swine/CH/IMH/03分离株能够导致实验猪出现神经症状,剖检可见脑、肾、脾、肠道发生出血,病理组织学检查可见脑实质内胶质细胞浸润、肾脏间质小血管淤血、脾淋巴细胞减少、小肠伴有出血及淋巴细胞浸润等病理变化,对仔猪具有强致病性。     

Prevalence and molecular characterization of porcine Picobirnavirus in piglets of North East Region of India

Tropical Animal Health and Production - Picobirnaviruses (PBVs) have been recognized as one of the important causal viral agents of gastroenteritis in several animal species especially in young...     

Phenotypic and genotypic characterization of Salmonella arizonae from an integrated turkey operation

Fifty cases submitted between 2000 and 2002 were selected for retrospective analysis to evaluate possible relationships between Salmonella arizonae isolated from breeder flocks, hatching eggs, and meat bird flocks belonging to a single turkey integrator. In all the meat bird cases selected for this study, arizonosis was the primary diagnosis. In birds under 1 month of age, clinical signs and pathologic changes were observed in older birds. The Salmonella arizonae isolates were analyzed by antibiotic resistance pattern and serotype and genotyped by pulsed-field gel electrophoresis (PFGE). Serotyping and PFGE yielded similar results, but the antibiotic resistance patterns did not correspond to either serotyping or PFGE typing. The presence of common pulsed-field patterns in breeder flocks, eggs, and meat bird flocks suggested that S. arizonae was being transmitted vertically from the breeder flock.     

猪捷申病毒3D蛋白的原核高效表达及其反应活性

根据猪捷申病毒（PTV）Swine／CH／IMH／03株核苷酸序列，设计了1对特异性引物，以全长基因组重组质粒pSK—PTVFL为模板扩增了3D基因，并将扩增产物定向克隆到原核表达载体pET30a（＋）中，阳性质粒转化BL21（DE3），阳性菌经0．3mmol／L IPTG诱导后，进行Western—blotting检测。结果显示，3D蛋白获得了高效表达，表达量占菌体总蛋白的66．1％，且重组蛋白能与PTV Swine／CH／IMH／03株阳性血清发生特异性反应。表明，3D蛋白能在大肠杆菌中高效表达，并具有良好的免疫原性，这为开发相应的鉴别诊断技术奠定了基础。