Construction of Recombinant Pseudorabies Virus Expressing Canine Distemper Virus H Gene and Analysis on Its Biological Characters

[Objective] The aim was to construct a recombinant pseudorabies virus expressing canine distemper virus H gene and investigate its biological characters.[Method] H gene of canine distemper virus（CDV）strain Onderstepoort was produced by RT-PCR,inserted into pcDNA3.1（＋）vector to construct a expression cassette,which was then subcloned into transfer vector p8AA,prior to the insertion of LacZ expression cassette.The resulting new transfer vector was named as p8AAZH.Subsequently,p8AAZH was co-transfected with the genome of pseudorabies virus（PRV）Bartha-K61 into BHK-21 cells to enable gene recombination and virus package,and the virus solution was collected as cytopathic effect occurring.A series of procedures including blue plaque purification,PCR identification,observation under electron microscope and Western blot were carried out to screen the recombinant pseudorabies virus and identify the protein expression of target gene.Meanwhile,growth curve of the recombinant virus was determined in BHK-21 cells.[Result] The H gene had been inserted into the genome of Bartha-K61 strain,and RPRV-H was the same as Bartha-K61 in the one-step growth curve and cytopathic effect in BHK-21 cells.[Conclusion] The recombinant pseudorabies virus was constructed,and the insertion of H gene did not influence proliferation of recombinant virus,which laid a foundation for development of recombinant canine distemper virus vaccine.

Preparation and Preliminary Identification of Fluorescein Labeled Monoclonal Antibody against Canine Distemper Virus

[Objective] The aim of the present study was to develop a direct immunofluorescence method for the diagnosis of canine distemper （CD） with FITC-conjugated monoclonal antibodies （FITC-McAb）.[ Metbod] The McAb against CDV, designated as CE3, was purified with protein G and labeled with FITC through agitation method. After purification and identification, the optimal working concentration of FITC-labeled CE3 was determined. Then 61 clinical samples of suspected canine distemper were detected by direct immunofluorescence assay. [ Result] The absorption test, blocking test and specificity test showed that the labeled antibody had high specificity and sensitivity, but didn＇t have cross reaction with canine parvovirus （CPV）, canine parainfluenza virus （CPIV）, canine adenovirus （CAV） and rabies virus （RV）. The optimal working concentration was 1：80. The positive rate of clinical suspected samples was 48%. [ Conclusion] The direct immunofluorescence assay developed in this study was rapid, specific and convenient, and had great significance for the early diagnosis of canine distemper.

ELISA Detection of Antibody against Canine Distemper Virus in Fox

Canine distemper is one of the most important common viral epidemics in farmed foxes. In this study, the whole blood samples of foxes that had been immunized with canine distemper vaccine were collected from a farming area in Changli County. The antibody against canine distemper virus in the 85 prepared serum samples were detected with a double-antibody sandwich ELtSA method. The results showed among the 85 samples from the 4 farms, 81 samples were antibody-positive, and only 4 samples had no antibody against canine distemper. The overall antibody level was higher in that farming area. It is indicated canine distemper vaccination and vaccination program have gotten attention from most of the local farmers, and the immune effect of canine distemper vaccine is better.

金鱼雌核发育单倍体胚胎血液循环障碍产生机制

为研究鱼类单倍体血液循环障碍产生机制,人工诱导获得金鱼(Carassius auratus)雌核发育单倍体胚胎并进行活体观察及邻联茴香胺染色,结果显示金鱼雌核发育单倍体胚胎存在不同程度的血液循环不良和红细胞生成缺陷。为进一步探讨其发生的分子机制,利用反义RNA整胚原位杂交技术比较分析了原始造血和血管发生关键基因scl(Stem cell leukemia)、gata-1(Gata binding protein-1)和flk-1(Fetal liver kinase-1)在雌核发育单倍体和自交二倍体胚胎中的表达模式。原始造血关键基因scl和gata-1的检测结果显示,在金鱼单倍体胚胎中,后侧板中胚层(Posterior lateral-plate mesoderm,PLM)不能正常迁移至胚体中线融合形成原始造血关键部位中间细胞群(Intermediate cell mass,ICM);同时,scl和gata-1的表达沿单倍体胚胎前后轴缩短,并在躯干区域逐渐减弱。血管内皮标记基因flk-1的检测结果显示,单倍体胚胎原始体轴血管结构紊乱,节间血管发育不良或者缺失,没有形成完整的血管系统。此外,活体观察及gsc(Goosecoid)和N-cadherin(Neural cadherin)原位杂交检测结果显示,金鱼雌核发育单倍体胚胎原肠胚期存在细胞运动缺陷。以上研究结果提示,细胞运动迁移异常导致ICM和原始体轴血管发育缺陷,同时红细胞分化减少及外周血管发育不全,可能是造成金鱼雌核发育单倍体胚胎血液循环障碍的重要原因。

犬瘟热病犬主要内脏器官的病理组织学变化

犬瘟热是由犬瘟热病毒（Canine distemper virus,CDV）引起的一种急性、热性、高度接触传染性、致死性传染病,病死率达36%-80%。犬瘟热对机体的损害是多系统多器官性的,临床上大多根据患犬的表观症状入手制定对应治疗方案,但对于一些无明显表观症状的病变器官没有针对性治疗,这亦是现今治疗方案的治愈率较低的原因之一。

Histopathological Observation of Canine Distemper in Raccoon Dogs

In this study, histopathological changes in the heart, liver, spleen, lung, kidney and other organs of raccoon dogs died of canine distemper were observed. According to the results, the lung of infected raccoon dogs exhibited severe hemor- rhage and emphysema; the liver was congested and degenerated; the spleen was enlarged; the exhibited severe hemorrhage and hyaline degeneration. Multiple organs of infected raccoon dogs were congested and hemorrhaged with tissue damage, in- flammatory cell infiltration and a series of pathological changes. This study laid a solid foundation for clinical diagnosis and treatment of canine distemper in raccoon dogs.

犬瘟热的诊断与防治

犬瘟热（Canine distemper,CD）是由犬瘟热病毒引起的犬科（尤其是幼犬）、鼬科及一部分浣熊科动物的一种急性、高度接触性传染病。以呈现双相热型、结膜炎、鼻炎、支气管炎、卡他性肺炎以及严重的胃肠炎和神经症状为特征。本病在毛皮动物养殖行业广泛流行,危害极大,造成巨大经济损失。

犬瘟热的诊断与防制

本文报导了长沙市某宠物医院就诊的100例CD患犬,其中呼吸型42例、消化道型30例、神经型4例、混合型24例。通过采取抗体疗法、血清疗法、对症疗法、输血疗法及中西医结合疗法等,取得了一定的治疗效果。其治愈率为35%,其中消化道型46.7%、呼吸道型35.7%、混合型25%、神经型0%。犬瘟热（Canine distemper CD）是由犬瘟热病毒（CDV）引起的犬和肉食目中许多动物的一种高度接触性传染病。

Canine Distemper Virus Utilizes Different Receptors to Infect Chicken Embryo Fibroblasts and Vero cells

Inducing animal viruses to adapt to chicken embryos or chicken embryo fibroblasts(CEF) is a common method to develop attenuated live vaccines with full security.Canine distemper virus(CDV) also does this,but the mechanisms and particular receptors remain unclear.Virus overlay protein blot assays were carried out on CEF membrane proteins,which were extracted respectively with a Mem-PER TM kit,a radioimmunoprecipitation assay buffer or a modified co-immunoprecipitation method,and revealed a common 57 kDa positive band that differed from the 42-kDa positive band in Vero cells and also from those receptors reported in lymphocytes and 293 cells,indicating a receptor diversity of CDV and the possibility of the 57-kDa protein acting as a receptor that is involved in adaptive infection of CDV Kunming strain to CEF.

水貂犬瘟热病毒RT-PCR检测方法建立及应用

水貂犬瘟热病毒（Canine distemper virus,CDV）属于副黏病毒科（Paramyxoviridae）麻疹病毒属（Morbillivirus）。形状多种,病毒直径大小多在150-300nm。本实验通过RTPCR技术检测了水貂病料中犬瘟热病毒的感染情况,建立具有快速、灵敏的RT-PCR检测方法。

犬瘟热的诊断与治疗

1病因 犬瘟热（Canine distemper,CD）是由犬瘟热病毒（Canine distemper virus,CDV）引起的犬科等动物的一种急性、高度接触性传染病。近几年,随着哈密市宠物犬养殖数量的增多,犬瘟热发病率也在升高。本病以预防为主,发病后应及时对症治疗。本文主要通过工作中所遇到的典型病例,从发病症状、诊断方法和治疗方法方面进行了详细的阐述,并就该病的相关内容进行了详细的讨论。

Ultrastructural features of PPRV infection in Vero cells

Dear Editor,The peste des petits ruminants virus(PPRV)causes an increasingly important viral disease of livestock that predominantly infects small ruminants such as goats and sheep.It belongs to the Paramyxoviridae family and is classified as the fourth member of the genus Morbillivirus because of its genetic similarity with other members of this genus,which includes measles virus(MV),rinderpest virus(RPV),canine distemper virus

实验犬暴发犬瘟热的诊疗

犬瘟热（Canine distemper,CD）俗称狗瘟,是由犬瘟热病毒（Canine distemper virus,CDV）引起的传染病,主要感染犬科和鼬科动物,具有高度接触性、传染性强、致病率高、流行速度快等特点。同时,该病还会引起一系列并发症,临床表现多样化,如支气管炎、肺炎、严重胃肠炎、腹泻和神经症状等特征,严重的会侵害动物的脑细胞,威胁犬的生命[1~3]。本病主要通过直接接触或近距离飞沫传播,

Pathogenesis of Demyelinating Encephalopathy in Dogs with Spontaneous Acute Canine Distemper

So far, the pathogenesis of demyelination caused by canine distemper virus （CDV） in the central nervous system has remained unclear, although a lot of studies have been done extensively. To further investigate the relation of variety cells in brain to demyelination, this study was performed on 15 dogs with spontaneous acute canine distemper and 2 controls. According to anatomical relation, the brain was divided into cerebrum, cerebral stem and cerebellum. The sections with no, mild, moderate, or severe demyelinating lesions were selected respectively and stained by HE and immunohistochemistry. Immuno-localisation of CDV antigen was used to conftrm CDV infection. The brain was examined for co-localisation of the CDV antigen with either an astrocyte-specific marker, glial fibrillary acidic protein （GFAP）, or an oligodendrocyte-specific marker, galactocerebroside （GalC）. Apoptotic cell was detected by TdT-mediated nick end-labeling assay （TUNEL）. The results demonstrated that the local disturbance of blood circulation mainly included congestion, edema, thrombosis, and disseminated intravascular coagulation （DIC）. The CDV neucleocapsid protein positive reaction, metabolic disorder and apoptosis of oligodendrocytes were observed in demyelinating areas. Lots of astrocytes displayed CDV antigen-positive, especially in their process. Some of them became apoptotic cell confirmed by TUNEL staining. Fibrous astrocytes showed more intense GFAP-positive in mild and moderate demyelinating area. Some of nervous cells located in pyramidal cell layers and nucleus nervi were in degeneration, necrosis. Satellitosis, neuronophagia and apoptotic neurons were examined by hematoxylin and eosin （HE） and TUNEL staining. The results suggested that the demyelinating changes in brain tissues infected with CDV mainly related to the metabolic disorder and apoptosis of ogliodendrocytes and astrocytes; also involved with the local disturbance of blood circulation and some neuron lost.

Pathomorphologic Study of Parasitic Enteritis Caused by Acute Canine Distemper

[Objective]The aim was to survey relationship between acute canine distemper and parasitic enteritis from pathology. [Method]Twelve cases of acute canine distemper with diarrhea were researched as per immunohistochemistry,Haematoxylin Eosin,and PAS staining kit. [Result] Of the twelve diseased dogs （ with diarrhea） ,six were detected caused by coccidium and two were detected by cryptosporidium. Coccidian protozoa is mainly in epithelial cells of jejunum and ileum,and some can be found in cut-off intestinal epithelial cells and in mucus formed by destroyed intesti- nal villus. The most common shapes of coccidian protozoa are trophozoite and schizont. The former is mainly within or among epithelial cells; nucle- us is in center and stained by hematoxylin; protoplasm is in ＂ fined mesh＂ shape. The latter,round or oval,contains much glycogenosome in de- generated intestinal epithelial cells. On the other hand,cryptosporidium is mainly in striated borders of intestinal epithelial cells and intestinal gland cells,leading to destruction of villus and cut-off of cells. Through detection on monoclonal antibody of nucleocapsid proteins of anti-canine distemper virus,it was found that epithelial cells in intestinal mucosa,glandular cells in recesses,lymphocytes and macrophage infittrated in lamina propria and dendritic cells in aggregated nodule were all with positive reactions. [Conclusion]Parasitic diarrhea caused by acute canine distemper occurs when resistance of intestinal mucosa caused by canine distemper virus begins to decline.

Construction of Recombinant Expression Plasmids Containing H and F Protein Genes of Canine Distemper Virus Isolated from a Mink and Their Expression in Prokaryotic Cells

[Objective] This study aimed to construct the recombinant expression plasmids containing H and F protein genes of Canine distemper virus isolated from a mink and to express these two genes in prekaryotic cells as well as to study the reactogenieity of the expressed products. [ Method ] RT-PCR amplification was used to obtain H and F protein genes; TA cloning and subclonlng techniques were used to construct the cloning plasmids（pMD-18T-H and pMD-18T-F） and recombinant expression plasmids（pET28a-H and pET28a-F） ; SDS-PAGE and Western-blotting were adopted to verify whether the target proteins were successfully expressed. [ Result] The recombinant expression plasmids pET28a-H and pET28a-F containing H and F protein genes of Canine distemper virus isolated from a mink were successfully constructed, and both the expressed H and F proteins with respectively relative molecular mass of 31 400 and 38 200 produced positive reac- tion with the CDV standard positive serum. [ Conclusion] The H and F proteins expressed in prokaryotic cells were the same with the natural ones in terms of reac- togenicity, which can be utilized for diagnosis of a CDV＇s infection or for an epidemiological investigation. Meanwhile, they also provide a basis for developing ge- netically engineered subunit vaccines.

犬瘟热及其防控措施

1病原体 犬瘟热病毒（Canine distemper virus CDV）属于副粘病毒科麻疹病毒属的成员,病毒粒子多呈球形,直径在150~300nm之间,病毒的基因组为负链RNA。圆形的病毒体内含有直径为15~17.5nm的螺旋形核心。病毒对乙醚和氯仿敏感。0.75%的福尔马林、1%的煤酚皂溶液或3%氢氧化钠溶液能在数小时内灭活病毒。CDV在p H4.5~9.0条件下均能存活。

First complete genomic sequence analysis of canine distemper virus in wild boar

Dear Editor,Canine distemper virus(CDV)is the causal agent of a highly contagious viral infectious disease that affects domestic and wild carnivores globally.It is an enveloped,non-segmented negative sense RNA virus that belongs to the Morbillivirus genus in Paramyxoviridae family,which contains viruses of epidemiological relevance to humans and animals.Based on the variability of the hemagglutinin gene(H),CDV strains have been at least 21 major genetic lineages:America-1 to America-5,Canada-1 and-2,Asia-1 to Asia-6,Europe wild-life,Arctic,Africa-1/South Africa,Africa-2,South America-1 to South America-3 and Rockborn-like(Giacinti et al.,2022).Mutations in the binding sites of the H protein,which interact with viral entry receptors such as signaling lymphocytic activation molecule(SLAM)and nectin-4,are associated with the emergence of the disease in new host species(Beineke et al.,2015).

犬瘟热的诊断和治疗

犬瘟热病毒（Canine distemper virus,CDV）的传染性极强,犬一旦有犬瘟热发生,无论采取怎样严密的防护措施,都不能避免同居一室的犬感染1-2。大量的临床实践经验表明,犬瘟热早期的治愈率高达98%以上,到了病程的中后期治疗就非常困难,特别是出现典型症状后再进行治疗则治愈率很低,死亡率较高。

消化道型犬瘟热病犬的胃肠道病理学观察

犬瘟热（canine distemper,CD）是由犬瘟热病毒（canine distemper virus,CDV）感染引起的一种高度接触性、致死性传染病,临床症状多样,容易继发感染其他细菌、病毒或寄生虫,死亡率高达30%-80%。