Experimental reproduction of viral chorioretinitis in kangaroos

OBJECTIVE: To investigate whether preparations containing Wallal and/or Warrego viruses could cause disease when inoculated subcutaneously into captive kangaroos. DESIGN AND PROCEDURE: Four groups of two kangaroos, seronegative to both Wallal and Warrego virus, were each inoculated with wild Wallal virus, cultured Wallal virus, wild Warrego virus, or wild Warrego virus followed by wild Wallal virus after 3 weeks. A single uninoculated animal served as a control. Animals were monitored weekly under anaesthesia, examined ophthalmoscopically (including fundic photography), and samples collected for haematological and serum biochemical analysis, virus isolation, PCR and serological examination for antibodies against Wallal and Warrego viruses. Animals inoculated with cultured Wallal virus were killed at week 10, and remaining kangaroos were reinoculated with cultured Wallal virus at week 12. RESULTS: Virus was isolated from the blood of two kangaroos 2 weeks after inoculation with Wallal virus preparations, and from a third kangaroo 2 weeks after reinoculation. By 3 weeks after inoculation, all kangaroos given Wallal virus preparations had seroconverted to Wallal virus and one had seroconverted to Warrego virus. Fundic changes were detected in the three viraemic kangaroos 4 or more weeks after inoculation, and lesions were present in the eye and brain typical of those seen in field cases of chorioretinitis. No other kangaroos had lesions. Wallal virus was identified by PCR and immunohistochemical analysis in the retina of one affected animal and orbivirus-like particles were seen by electron microscopy in the remains of retinal cells. CONCLUSION: The condition of chorioretinitis was reproduced in three of eight kangaroos by inoculation with preparations containing Wallal virus.     

Updated distribution and host records for the argasid tick Ornithodoros (Pavlovskyella) zumpti: A potential vector of African swine fever virus in South Africa

African swine fever virus (ASFV) causes a lethal and contagious disease of domestic pigs. In South Africa, the virus historically circulated in warthogs and ornithodorid ticks that were only found in warthog burrows in the north of the country. Regulations implemented in 1935 to prevent transfer of infected animals or products to the south initially proved effective but from 2016 there have been outbreaks of disease in the south that cannot be traced to transfer of infection from the north. From 1963 there were widespread translocations of warthogs to the south, initially from a source considered to be free of ornithodorid ticks. We undertook to determine whether sylvatic circulation of ASFV occurs in the south, including identification of potential new vectors, through testing extralimital warthogs for antibody and ticks for virus. Results of testing warthogs for antibody and other species of ticks for virus will be presented separately. Here we report finding Ornithodoros (Pavlovskyella) zumpti ticks in warthog burrows for the first time. This occurred in the Eastern Cape Province (ECP) in 2019. Since African swine fever was recognised in the ECP for the first time in 2020 and outbreaks of the disease in domestic pigs continue to occur there, priority should be given to determining the distribution range and vector potential of O. (P.) zumpti for ASFV.     

干扰素诱导跨膜蛋白1对猪源冠状病毒吸附宿主细胞的影响

为探索干扰素诱导跨膜蛋白1（interferon-inducible transmembrane protein 1,IFITM1）对猪源冠状病毒复制的影响,本研究选用猪传染性胃肠炎病毒（Transmissible gastroenteritis virus,TGEV）高致病毒株及其宿主细胞PK15作为研究对象。正常PK15细胞接种TGEV后,实时荧光定量PCR检测感染后不同时间点PK15细胞中一些干扰素刺激基因（interferon-stimulated genes,ISGs）的mRNA表达水平;利用慢病毒表达系统构建稳定表达和稳定干扰IFITM1表达的PK15细胞系。将一段靶向干扰猪源IFITM1的shRNA序列及猪源IFITM1全长,分别插入pLKO.1-EGFP-Puro载体及pLVML-Myc-MCS-IRES-Puro载体中,分别构建出pLKO.1-IFITM1shRNA-EGFP-Puro及pLVML-Myc-IFITM1-IRES-Puro重组质粒。将重组质粒与慢病毒包装质粒共转染293FT细胞后获得带有目的基因的重组慢病毒,慢病毒侵染PK15细胞后用嘌呤霉素进行筛选,获得稳定表达及稳定干扰IFITM1表达的PK15细胞系,分别命名为PK15-IFITM1及PK15-IFITM1^-/-,并分别用实时荧光定量PCR、间接免疫荧光试验（IFA）及Western blotting检测IFITM1干扰效率及表达情况;TGEV接种PK15-IFITM1^-/-和PK15-IFITM1,实时荧光定量PCR测定细胞中TGEV的拷贝数。结果显示,PK15细胞接种TGEV后的48 h内,一些ISGs的mRNA水平均有所上升;PK15-IFITM1^-/-细胞系的干扰效率为70%,PK15-IFITM1细胞系表达成功;在PK15-IFITM1^-/-细胞系中,IFITM1的mRNA水平显著下调,促进了TGEV的复制。反之,在PK15-IFITM1细胞系中,TGEV的复制受到了抑制。但IFITM1的表达或缺失却不影响TGEV对PK15的吸附作用。总之,IFITM1对TGEV有显著的抗病毒作用,IFITM1不影响TGEV对PK15细胞的早期吸附,这为后续IFITM1抗冠状病毒机制的研究奠定了基础。     

Study on Synergistic Effect of VA5 Immunopotentiator on Pigeon NDV Inactivated Vaccine

This study was designed to evaluate the synergistic effect of VA5 immunopotentiator on pigeon ND4416 inactivated vaccine.160 healthy pigeons were randomly divided into four groups,including ND4416 strain inactivated vaccine group (NDV group),ND4416 strain inactivated vaccine and immunopotentiator (VA5) mixed group (NDV+VA5 group),La Sota inactivated vaccine group (La Sota group) and normal saline as the control group (C group),to assess their vaccine efficacy against virulent pigeon NDV by serological analysis and animal testing.Pigeons sera were collected at different time points after immunization and measured the HI antibody titer of each group.The results showed that VA5 immunopotentiator significantly improved the serum antibody level of pigeons immunized with pigeon Newcastle disease inactivated vaccine (P<0.05).In addition,comparative test of spleen lymphocyte transformation was conducted at various time points after immunization.The results indicated that VA5 effectively stimulated the lymphocyte transformation of immune pigeon.Pigeons in each groups were challenged with ND4416 strain at the 30th,90th and 180th d after immunization.The results presented that the NDV+VA5 group had 100% protection rate and higher than La Sota group.The duration of immunization test showed that the antibody titer of NDV+VA5 group reached peak 11.20log₂ at the 21st d,remained 7.50log₂ at 180th d,and the protection rate remained 100% at 180th d.It indicated that VA5 immunopotentiator sustained the immune duration of pigeon NDV vaccine up to 180 d.Moreover,the in vitro detoxification test results suggested that VA5 immunopotentiator reduced the in vitro detoxification cycle of pigeons after challenge.Overall,this study suggested VA5 immunopotentiator could significantly improve the immune efficacy of pigeon Newcastle disease inactivated vaccine,which provided a basis for further enhancing the efficacy of Newcastle disease vaccine,as well increased experimental data for the application of immunopotentiators.     

肾型传染性支气管炎病毒对鸡嘌呤代谢的影响研究

用T株肾型传染性支气管炎病毒(IBV)人工感染14日龄非免疫土种鸡建立感染模型,于攻毒前、攻毒后48 h、72 h、96 h、120 h 采集血样,测定血浆黄嘌呤氧化酶(xanthine oxidase, XOD)活性,探索鸡嘌呤代谢变化与肾型IBV致病机理之间的关系.结果表明,与攻毒前相比较, 攻毒后48 h和72 h血浆XOD活性明显增强, 差异显著(P<0.01,P<0.05).结果提示,XOD活性升高引起的嘌呤代谢紊乱可能是鸡肾型IB的病理机制之一.     

表达H3N2亚型猪流感病毒HA基因重组伪狂犬病病毒的构建

将SV40启动子控制下的LacZ基因表达盒和CMV启动子控制下的H3N2亚型猪流感病毒（SIV H3N2）的HA基因插入到伪狂犬病病毒（PRV）通用转移载体pBdTK-Uni中，获得转移载体pLTK-HA。将该载体与PRV Bartha-K61株基因组DNA通过脂质体法共转染Vero细胞，经过10代蓝斑筛选、纯化和PCR鉴定获得了一株插入SIV HA基因的重组伪狂犬病病毒，命名为rPRV-HA。Western blotting和间接免疫荧光试验证实HA基因在重组病毒感染的细胞中获得了表达。用不同的细胞（PK-15、IBRS-2、Vero和鸡胚成纤维细胞）对该重组病毒与亲本病毒的增殖滴度和致细胞病变进行比较，未见显著差异，对第30代重组病毒的HA基因进行序列分析，表明该重组病毒遗传性状稳定。     

甲型肝炎病毒免疫控制研究进展

对甲型肝炎病毒(HAV)发病机理和免疫调控的研究近年来重新受到关注。在发展中国家感染者的平均年龄增加,导致机制尚不明确的更严重的肝炎。而且,从HAV和丙型肝炎病毒(HCV)的免疫对比来看,这两种正链RNA病毒感染结果完全不同。HCV比HAV复制效率低,导致HCV蛋白表达量低,因此对IFN信号传导的拮抗作用较低。有研究发现循环的HAV病毒颗粒隐藏在膜里,导致先天免疫和抗体介导中和作用的激活。同时还考虑到CD4^+辅助T细胞对CD8^+细胞毒性T细胞对抗病毒免疫和肝损伤的作用,提出了一种非细胞毒性的HAV感染免疫控制模型。     

鹿源狂犬病野毒8202株基因型的研究

为了从基因水平确定鹿源狂犬病野毒8202株与其它狂犬病病毒的进化关系,应用RT-半嵌套式PCR技术,利用3条引物对病毒的核蛋白基因进行扩增、克隆及序列测定,并与已发表的狂犬病病毒株3aG、PV、CVS、HEP-Flury、RC-HL、Nishigahara、SADB19的核蛋白基因进行了比较分析。结果表明,8202株与其它7个病毒株均来源于同一个进化枝,属于基因Ⅰ型。     

An oculo-cerebellar syndrome caused by congenital bovine viral diarrhoea virus-infection

An epizootic characterized by birth of calves severly ataxic and blind were encountered in 3 herds 7–8 months after outbreaks of bovine virus diarrhoea. Serological and virological investigations indicated introduction of bovine viral diarrhoea virus (BVDV) into previously virus-free herds, followed by transplacental virus infection of the fetuses of cows in the first trimester. Clinical, pathological, serological, and microbiological examinations were performed on 10 calves. Pathological findings included microcephaly and cerebellar hypoplasia, ocular malformations, and thymic hypoplasia. BVDV was isolated from tissue and blood of 7 calves, and 4 calves, 1 of which had not received colostrum, had virus-specific neutralizing antibodies.This is the first report on natural occurrence of congenital bovine infection with BVDV among Danish cattle herds resulting in abortion and birth of calves with severe debilitating congenital anomalies. It draws attention to the importance of this virus for bovines of all age groups.     

Comparison of antibody values in sera of pigs vaccinated with a subunit or an attenuated vaccine against classical swine fever

Ten pigs, aged 85 days, were vaccinated with a subunit vaccine containing 32 g of classical swine fever virus glycoprotein E2 (gp E2) (group 1), and a further 10 pigs were vaccinated with a C strain vaccine (10^4±0.15 TCID₅₀/ml), produced by amplification in minipig kidney (MPK) cell culture (group 2). Nine non-vaccinated pigs served as a control group (group 3). Serum samples were collected before (day 0) and at 4, 10, 21 and 28 days after vaccination and were analysed by two commercially available enzyme immunoassays and by a neutralizing peroxidase-linked assay (NPLA). At the same times, peripheral blood was taken for determining the total leukocyte count and the body temperature was taken daily. Antibodies were not detected in serum samples collected before vaccination (day 0), and no side-effects that could be connected with vaccination were observed during the trial. Ten days after vaccination 6/10 pigs vaccinated with the subunit vaccine were seropositive. On days 21 and 28, the ratios of serologically positive to vaccinated pigs were 9/10 and 10/10, respectively. Four of the ten pigs that were vaccinated with the C strain vaccine were positive on day 21 and 9/10 on day 28. However, the results of the NPLA showed that only 4/10 pigs had an antibody titre >1:32 at the end of the trial in both the vaccinated groups, even though the subunit vaccine initiated an earlier and higher level of neutralizing antibodies than the vaccine produced from the C strain. Challenge was performed 28 days after vaccination on four randomly selected pigs from both vaccinated groups. The pigs survived the challenge without showing any clinical signs of classical swine fever (CSF), while two nonvaccinated control pigs died on the 10th and 12th days after infection.