An investigation into the basic virus-antibody neutralization reaction, with special regard to the reaction in the constant-virus/varying-serum neutralization test.

A study of the basic reaction in neutralization of virus (V) by virus-neutralizing antibody (VNA) was performed with infectious bovine rhinotracheitis virus and serum collected from naturally and experimentally infected cattle after the primary immunization phase. In constant-virus/varying-serum neutralization tests a direct proportionality between VNA titer and length of preincubation was observed and found to be in accordance with basic laws of neutralization. A deviation from this direct proportionality, which was partly attributed to the presence of a dissociable V-VNA complex, was seen with relatively short preincubation. Expressing a relationship between VNA titer, length of preincubation, and virus dose under conditions where a dissociable V-VNA complex can be ignored, a log. VNA/log. V equivalence factor of neutralization was introduced. A linear relationship was found between VNA titer, taken logarithmically, and preincubation temperature. A rise in temperature by 10°C gave an increase in VNA titer of approx. 1.2 in log₂. Formulae are presented for the neutralization rate factor corrected for a demonstrated invalidity of the percentage law, and for the relation between the neutralization rate factor and VNA titer. It is concluded that the results presented have elucidated the possibilities of improving the sensitivity of neutralization tests.     

Bluetongue Virus （BTV） Serological Survey and Evidence of Emergent BTV-8 Serotype in Morocco

Bluetongue （BT） is an infectious, non-contagious arthropod-borne disease that infects all ruminants, including sheep, cattle, deer, goats and camelids. Bluetongue virus （BTV） belonged to Reoviridae is ARN genome of 19 Kb. Twenty-six BTV serotypes have long been recognized, to be associated with severe disease in certain breeds of sheep, whereas cattle and goats are usually sub-clinically affected. Before 2004, BT was considered an exotic disease in Morocco, however, the first outbreak was observed in 2004 in sheep. This outbreak was caused by the isolated BTV-4. Two years later a BTV-1 emerged in Morocco. Both serotype 1 and serotype 4 circulated after 2007 across the country. The aims of the present work was to perform a serological study on sheep from different regions in Morocco in order to clarify the current BTV epidemiological situation and its evolution from 2009 to 2012, to determine the co-infection rate, and to confirm the possible circulation of other BTV Serotype mainly the BTV-8. All of 436 sera were tested by serum neutralization using reference strains. Results confirm the presence of BTV-4, BTV-1 and BTV-8. However, the present study report for the first time the emerging BTV-8 circulation in Morocco. Moreover, the founding reveal as well a higher co-infection rate in cattle compared to sheep.     

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.     

A rapid immunoperoxidase-based virus neutralization assay for salmonid alphavirus used for a serological survey in Northern Ireland

A modified virus neutralization (VN) assay was developed to replace an existing assay read on the presence or absence of virus-induced cytopathic effect (CPE). The modified assay used a monoclonal antibody to salmon pancreas disease virus as the first layer of an immunoperoxidase (IPX)-based immunostaining technique to detect viral growth. The IPX-based VN assay required only 3 days to perform, and the adoption of a 96-well microtitre format facilitated a high throughput of samples requiring small volumes of serum, cells and virus. When 352 sera from farmed salmon and 302 sera from farmed trout were tested by both the modified and the original CPE-based assays, overall correlations of 97.72 and 96.03% were, respectively, obtained (96.94% combined). When the modified assay was used to test 188 sera collected from wild salmonids in freshwater river systems in Northern Ireland, no positive results were recorded.     

犬瘟热病毒抗体检测方法的比较研究

用犬瘟热（ＣＤ）弱毒和自制的高免犬血清，建立了检测犬瘟热病毒（ＣＤＶ）抗体的中和（ＳＮ）试验、间接荧光抗体技术（ＩＦＡＴ）和间接免疫酶染色法。这３种方法对３８份ＣＤ弱毒疫苗免疫犬血清抗体效价的测定结果表明，当被检血清稀释度为１∶１００时，ＳＮ试验、ＩＦＡＴ和间接免疫酶染色法测定时的阳性血清数分别为２９，７和３１份，ＩＦＡＴ和间接免疫酶染色法测定结果相对于ＳＮ试验结果的符合率分别为４２．１％和９４．７％。同时发现，当ＳＮ试验测定的血清效价在１∶１００以上时，ＩＦＡＴ测定结果总是在１∶２５以上，间接免疫酶染色法测定结果总是在１∶１００以上。３种方法以各自初步确定的ＣＤＶ血清抗体效价完全保护值指标计算所测血清的完全保护率，ＳＮ试验为７６．３％，ＩＦＡＴ为７８．９％，间接免疫酶染色法为８１．６％。据此认为，ＩＦＡＴ和间接免疫酶染色法均可部分代替ＳＮ试验用于ＣＤ疫苗免疫效果的评价、免疫犬抗体水平的监测以及ＣＤ的流行病学调查     

徐州地区牛流行热病毒的分离与鉴定

1991年夏秋之交徐州市各奶牛场暴发牛流行热。为确诊该病,采集了发病高热期抗凝血,进行了病毒分离和鉴定试验。病料经处理后,直接接种于BHK_21细胞,传至第二代可见典型的CPE。电镜下观察到80—150nm弹状病毒颗粒。分离毒的理化特性、核酸类型及中和试验结果均与标准BEFV相一致,证实该分离病毒为牛流行热病毒。     

A comparative examination of swine sera for antibody to Aujeszky virus with the conventional and a modified virus-serum neutralization test and a modified direct complement fixation test.

Sera of pigs from élite breeding herds, of boars and sows collected at slaughter-houses, and of pigs from herds known to be infected, were examined for antibody to Aujeszky virus. The conventional and a modified virus-neutralizing antibody (VNA) test and a modified direct complement fixation (CF) test were employed. In simultaneous titrations of positive sera the modified VNA test gave titers approx. 4 log2 units above the titers obtained by the conventional test. The conventional VNA test was found insufficiently sensitive. Unspecific neutralization in the modified VNA test was infrequent in serum dilution 1/2 and rare in dilution 1/4. The GF tests on sera of slaughter sows and animals from known infected herds showed a remarkable consistency with the VNA tests. Inconsistent results were obtained with but few sera. Abt. 5 % of the sera could not be examined because of complement fixation with control antigen.     

小反刍兽疫病毒N蛋白抗体与H蛋白抗体在羊体内代谢消长规律的研究

为研究小反刍兽疫病毒N蛋白抗体与H蛋白抗体在羊体内的代谢消长规律,试验通过建立小反刍兽疫N蛋白双抗原夹心ELISA抗体检测方法、H蛋白阻断ELISA抗体检测方法与血清中和抗体试验方法,分别检测了羊免疫疫苗后体内N蛋白抗体与H蛋白抗体在每个免疫阶段的代谢消长变化。结果表明:羊免疫小反刍兽疫疫苗后,血清内的N蛋白抗体与H蛋白抗体在6~8周时血清抗体效价较高,对应的羊体内中和抗体效价为1∶512,且效价至少可以持续到10周以上,2种抗体具有一定消长代谢规律的相关性。该研究结果也为以N抗原与H抗原为基础建立相应的抗原捕获ELISA方法、竞争ELISA检测方法、间接ELISA检测方法以及双抗原夹心ELISA检测方法奠定了一定的理论依据。