Safety and efficacy of an experimental reovirus vaccine for in ovo administration

A commercial reovirus vaccine alone or experimental reovirus vaccine plus antibody complex were inoculated into 18-day-old specific pathogen free (SPF) broiler embryos at 0.1 of the recommended chick dose. The following groups were used: group 1A was not vaccinated or challenged; group 1B was not vaccinated, but was challenged with virulent reovirus; group 2 received the vaccine complexed with 1/4 dilution of antiserum; group 3 received the vaccine with 1/8 dilution of antiserum; group 4 received the vaccine with 1/16 dilution of antiserum, and group 5 received vaccine alone. At 1, 3, 6, 9, and 12 days of age, serum was collected and antibody against avian reovirus was analyzed by enzyme-linked immunosorbent assay (ELISA). At the same times, spleens were collected and vaccine virus detected by inoculating chicken embryo fibroblasts (CEFs) and examining for cytopathic effect. At 15 days of age, chickens in groups 2-5 were challenged with reovirus. At 22 days of age, birds were euthanatized and weighed. Efficacy of the vaccines was based on safety, percent protection, and antibody response. In ovo vaccination with the commercial or experimental vaccines did not adversely affect hatchability of SPF chickens. The vaccine complexed with antibody resulted in significantly less posthatch mortality (3.7%) when compared to mortality of chickens that received vaccine alone (17%). Both vaccine virus recovery and antibody response were delayed at least 3 days in birds receiving the experimental vaccines. In evo administration of reovirus antibody complex vaccines provided at least 70% protection. The experimental reovirus-antibody complex vaccines were safe and efficacious when given in ovo to SPF broiler embryos.     

犬呼肠病毒3型在BHK-21细胞上增殖的研究

采用酶标ＳＰＡ法进行了犬呼肠３ 型病毒在ＢＨＫ２１ 细胞上定位的研究。结果表明：接毒后６ ｈ 即可在胞浆见到絮状阳性反应,２４ ｈ 后呈强阳性着染;Ｈ．Ｅ．染色在胞浆内可见强嗜酸性着染或嗜酸性包涵体,提示可能是病毒感染引起的特征性病变,感染后细胞病变发展较快,６０ ｈ 细胞大部脱落;病毒血凝滴度结果也证实了细胞病变与病毒增殖有关。     

3种禽类呼肠孤病毒血清学相关性及致细胞病变差异分析

本研究旨在研究3种不同疾病型禽类呼肠孤病毒间的抗原性关系及病毒的培养特性。作者通过血清中和试验测定了禽呼肠孤病毒(ARV S1133株)、番鸭呼肠孤病毒(MDRV 9710株)、新型鸭呼肠孤病毒(NDRV NP01株)3种禽类呼肠孤病毒的血清学相关性,统计抗原相关性R值;并应用部分禽胚原代细胞及哺乳动物传代细胞对这3种病毒的培养特性进行了初步研究。结果表明,3种病毒株之间的R值很小,抗原相关性较低;三者具有广泛的细胞亲嗜性,能在多种细胞中增殖,并产生细胞病变,但病毒致细胞病变特征有所差异,ARV和NDRV均以巨融合为主,而MDRV则以细胞圆缩坏死为主。上述结果表明导致禽类不同疾病的ARV、MDRV和NDRV三者之间的抗原相关性较低,病毒的细胞培养特性也不同,细胞病变类型的差别提供了一种初步鉴别禽类呼肠孤病毒的方法。     

The propagation of avian viruses in a continuous cell line (QT35) of Japanese quail origin

Seven of nine avian virus families tested (Birnaviridae, Coronaviridae, Herpesviridae, Paramyxoviridae, Poxviridae, Reoviridae, and Retroviridae) were found to replicate in a quail fibroblast cell line, designated QT35, resulting in a cytopathic effect (CPE) visible with the naked eye or by low-power microscopy. In comparison, only one (Paramyxoviridae) of seven mammalian virus families tested produced an observable CPE. Cytopathic changes induced by examined viruses were round cell, syncytial, and focus formation. Trypsin did not promote cytopathic changes by selected CPE-negative avian and mammalian viruses in QT35 cells. Several avian viruses (infectious bursal disease virus, Newcastle disease virus, Canary pox virus, and reovirus) formed plaques under agar. Avian reovirus and infectious bursal disease virus produced similar titers in chicken embryo fibroblast (CEF) and QT35 cell cultures. Chicken-egg-yolk neutralizing-antibody titers to IBDV were comparable in CEF and QT35 cell-culture systems.     

A field trial with a multicomponent inactivated respiratory viral vaccine.

An inactivated virus vaccine containing strains of parainfluenza type 3 (PI3), bovine adenovirus type 3, reovirus type 1, bovine virus diarrhoea (BVD) and infectious bovine rhinotracheitis (IBR) viruses was tested in a group of 58 calves reared in a semi-intensive management system. Following vaccination, 1/30, 14/30 and 17/30, showed significant rises in antibody titre to reovirus type 1, adenovirus type 3 and IBR respectively. None of the animals showed significant serological response to PI 3 and BVD. In the control group, 2/28, 1/28, 6/28 and 3/28 developed antibody responses to reovirus type 1, BVD, adenovirus type 3 and IBR respectively. Microbiological examination revealed the presence of a wide variety of commensal bacteria and Mycoplasma bovirhinis in both groups. Analysis of the records of clinical examinations indicated that the respiratory tract infections occurred among the calves at between 50 and 80 days after arrival at the farm, and that there was no significant difference between the test and the control groups. A number of animals had maternal antibodies to the various components of the vaccine present before the trial commenced and these antibodies appeared to interfere with the subsequent serological response to the antigen challenge. The vaccination schedule recommended by the manufacturer does not entirely circumvent this problem.     

Efficacy of coarse-spray administration of a reovirus vaccine in young chickens.

Coarse-spray (CS) administration of a commercial S1133 reovirus vaccine in chickens for prevention of clinical viral tenosynovitis (VT) infection was evaluated. In Expt. 1, one-day-old specific-pathogen-free (SPF) white leghorns were vaccinated with a combination of reovirus, Newcastle disease (ND), and infectious bronchitis (IB) vaccines by CS and infectious bursal disease vaccine by the subcutaneous (SQ) route. In Expt. 2, one-day-old commercial broilers were vaccinated by CS with reovirus vaccine and Marek's disease (MD) vaccine by SQ. In Expt. 3, one-day-old commercial broilers received reovirus vaccine in combination with ND-IB vaccines at 1 day of age by CS and MD vaccine by SQ. Some birds received an initial or second vaccination at 7 days of age by CS or the drinking-water (DW) route. Birds vaccinated by CS at 1 day of age with reovirus vaccine did not produce circulating virus-neutralizing antibody against reovirus, although they had resistance to VT infection. In contrast, initial or booster vaccination at 7 days of age by CS or DW resulted in an antibody response and greater resistance to challenge than did CS vaccination at 1 day of age. There was no difference in efficacy between CS and DW routes at 7 days of age. The reovirus vaccine did not interfere with other vaccines as measured by serologic (ND-IB-IBD) or challenge (MD) studies.     

Immune response to avian reovirus in chickens and protection against experimental infection

Objectives To assess the efficacy of the vaccination procedure and the effect of the transfer of maternal antibodies to progeny chickens on reovirus pathogenicity.
Design To vaccinate chickens and challenge progeny chickens with high doses of homologous and heterologous viruses.
Procedure High doses of reovirus strains RAM-1, 1091 and 724 were used to induce tenosynovitis lesions. High doses were produced by concentration of viruses grown in cell culture. Then similar doses of viruses were used to challenge immunised chickens progeny.
Result Vaccination of breeding hens with the RAM-1 strain of avian reovirus, which resulted in the passive transfer of neutralising antibody to progeny chickens, completely prevented the development of tenosynovitis in 80% of progeny chickens infected with the homologous virus. Even though multiple injection of hens resulted in broadening of the normal type-specificity of the neutralising antibody response against heterologous serotypes of avian reovirus, only marginal protection against strains of two heterologous serotypes of avian reovirus was obtained.
Conclusions A model for assessing the efficacy of vaccination against avian reovirus strains on clinical signs such as tenosynovitis was developed that overcome the normal low virulence of Australian strains of avian reovirus. Breeding hens can be immunised with Australian strain of avian reovirus with passive transfer of antibody via the yolk to the progeny chickens. Although the neutralising antibody response to three injections of inactivated virus decreased the specificity of the neutralising antibody response against antigenically heterologous strains of avian reovirus, the protective immunity appeared to retain type-specificity.     

Single and combined infections of specific-pathogen-free chickens with infectious bursal disease virus and an intestinal isolate of reovirus

The susceptibility of 1-day-old and 7-day-old specific-pathogen-free chickens to infection with a virulent strain of infectious bursal disease virus (IBDV) or an intestinal isolate of avian reovirus, or a combination of the two, was investigated. Chickens infected with IBDV and reovirus had more severe pathological lesions than chickens infected with either virus alone, and prior infection with IBDV enhanced the pathogenicity of enteric reovirus. Virus recovery was attempted from bursa, spleen, thymus, liver, intestine, pancreas, cecal tonsils, heart, and tarso-metatarsal tendons. Viruses were recovered from all tissues sampled for either IBDV or reovirus isolation, and indications were that infection with IBDV before infection with reovirus led to longer persistence of reovirus in some tissues. Antibodies to IBDV or reovirus were measured by the virus neutralization test and enzyme-linked immunosorbent assay. Chickens infected with IBDV had lower (P less than 0.05) antibody responses to reovirus than chickens infected with reovirus alone.     

Reovirus type 2 in domestic cats: isolation and experimental transmission

Five reovirus isolates were recovered in MA104 cell cultures from the faeces of three cats with nictitating membrane protrusion and diarrhoea, one cat with diarrhoea only and from one healthy cat. Four of these isolates were characterised as reovirus type 2 and one as reovirus type 3 by haemagglutination-inhibition and serum neutralization tests. Reovirus type 2 has not been reported previously in cats. Mild clinical signs of diarrhoea were noted in kittens infected experimentally with one of the feline reovirus type 2 isolates.     

Isolation of reoviruses from mink]

In this first report of the isolation of reovirus from mink, isolates were obtained from 18 to 26 young mink with viral enteritis, by using cultures of cat kidney cells. The isolated also produced cytopathic changes in cell cultures of mink kidney, dog kidney, piglet kidney, calf kidney, bovine embryonic kidney and calf testis. A characteristic feature was the formation of eosinophilic inclusion bodies in the cytoplasma of culture cells. Haemagglutination tests were negative with erythrocytes from cat, rabbit, pig, horse and cattle. Attempts to infect old and young mink, kittens and ferrets with tissue culture material failed. It was not known to what extent this second infection with reovirus influenced the course of mink viral enteritis.     

Monitoring the Immune Status of Broilers Against Reoviruses Using Challenge and Serologic Data

Reoviruses are an important cause of suboptimum performance in commercial broilers worldwide. Integrators use the enzyme-linked immunosorbent assay against the S1133 antigen for monitoring serum of breeders for indicating pullet vaccine success. However, without correlating serology to reovirus challenge, it is difficult to determine whether titers reflect protective immunity. We developed a broiler challenge test against 2 common reovirus isolates (2408 and S1133) to evaluate the efficacy of reovirus pullet vaccine programs. Two reovirus serologic and challenge studies were undertaken using chicks from broiler integrators from the southeastern United States. Breeder flocks, from which the chicks were obtained, received at least 1 live and 2 inactivated reovirus vaccines during their pullet phase. One-day-old progeny were collected from 6 breeder flocks. At 1 d of age, 20 chicks from each broiler flock were bled, and serum was analyzed for antibodies. At 3 to 4 d of age, 20 progeny per flock were challenged with the 2408 reovirus by intratracheal route. At 10 to 14 d of age, another 20 birds per flock were challenged with the S1133 reovirus by footpad. Twenty birds per flock were used as nonchallenged controls. At 3 wk of age, all birds were killed and weighed. Percentage of protection was calculated for each flock based on the absence of gross lesions. Flocks with at least 50% protection were considered well protected. Most flocks were well protected against both viruses. The percentage of protection correlated with day-old enzyme-linked immunosorbent assay titers. Chicks from younger hens had higher titers and the best protection against challenge. Producers, whose hen flocks were monitored herein, were doing a good job of immunizing pullets against reovirus. They are now using reovirus progeny challenge studies along with breeder antibody titers to determine vaccination success of their pullets.     

Serological response and virus shedding of chickens inoculated with reovirus via different routes

A comparison of the serological responses to a reovirus administered by the eye-drop, intramuscular, oral and subcutaneous routes was made in three-week-old chickens. Their patterns of virus shedding were also compared. All four routes resulted in an antibody response and no virus excretion in faeces more than two to three weeks after administration. The oral route represented the most favourable route for live reovirus vaccine administration.     

Serological Incidence of Avian Reovirus Infection in Broiler-breeders and Progeny in Nova Scotia

The plaque neutralization test and the agar gel precipitation test were used to detect neutralizing and precipitating antibody to avian reovirus strain WVU2937 in sera from 14 commercial broiler breeder flocks and eight progeny flocks. Ten breeder flocks (71%) possessed positive agar gel precipitation reactors (598 sera tested) and 12 (86%) possessed plaque neutralization reactors (114 serum pools tested). All broiler flocks possessed agar gel precipitation reactors, but these were not examined for neutralizing antibody. As no avian reovirus vaccine had been used in Canada prior to this survey these reactions were presumably due to natural infection. Clinical evidence suggested that such infection was asymptomatic in most cases.The development and persistence of antibody to reovirus strain WVU2937 was followed in chicks exposed to the virus by oral inoculation or by contact. Neutralizing antibody was demonstrated in all birds by postinoculation day 17 and this persisted for at least six months. Precipitating antibody was also demonstrated in most chicks by postinoculation day 17, but by six months only 12% of the chicks were still agar gel precipitation positive.     

Antigenic comparisons of selected avian reoviruses by use of the plaque-reduction neutralization assay

The antigenic interrelatedness of 3 clone-purified turkey reoviruses (NG-Turkey, 82-88, and NC-TEV) to each other and to 4 clone-purified chicken reoviruses (S1133, Co8, Fahey-Crawley, and avian type 2) was determined in reciprocal cross-neutralization tests, using polyclonal antisera and the plaque-reduction technique. The morphologic features of plaques formed under agar were studied for all 7 reoviruses, and size comparisons for turkey vs chicken isolates were made. All 3 turkey reoviruses (with the exception of NG-Turkey vs Fahey-Crawley chicken reovirus) formed plaques significantly (P less than 0.05) smaller than plaques produced by their chicken counterparts. The 3 turkey reoviruses were closely related to each other and to chicken reovirus CO8. The antigenic differences between turkey reoviruses 82-88 and NC-TEV and chicken reovirus S1133 were slight (minor subtype); however, the latter and NG-Turkey were serotypically distinct. The NG-Turkey and 82-88 turkey reoviruses were more related (minor subtype) to the Fahey-Crawley and avian type 2 chicken reoviruses, than was NC-TEV turkey reovirus (major subtype).     

Effect of in ovo administered reovirus vaccines on immune responses of specific-pathogen-free chickens

We investigated the effect of in ovo administered reovirus vaccines on the immune responses of specific-pathogen-free chickens. T-cell mitogenic responses to concanavalin A were numerically lower at 9 and 12 days of age and significantly lower at 6 days of age in birds vaccinated with a commercial reovirus vaccine compared with unvaccinated birds or birds vaccinated with an experimental reovirus-antibody complex vaccine. There were no significant differences in proportions of subpopulations of helper (CD4+CD8-) or cytotoxic (CD4-CD8+) T cells except at 12 days of age, when the percentages of CD4-CD8+ cells in the two vaccinated groups were statistically higher than in the nonvaccinated group. B-cell populations were not different among vaccine groups except at 9 days of age, when the vaccinated groups had the highest level of B cells. This commercial reovirus vaccine should not be given in ovo to embryos having little or no maternal antibody, otherwise immunosuppression may occur in the chicks. The addition of the antibody complex to the vaccine prevented this T-cell immunosuppression.     

Cell culture propagation of porcine rotavirus (reovirus-like agent).

Two isolates of porcine rotavirus (reovirus-like agent) were isolated and passaged in primary procine kidney cell cultures. Viral infectivity for cells was monitored by immunofluorescence because viral cytopathic effect was moderate. Successful passage of virus in cell culture required that viral suspensions obtained from infected cell cultures be treated with pancreatin prior to inoculation onto cell monolayers. Porcine rotavirus passage in cell culture also was accomplished, using trypsin treatments in lieu of pancreatin treatments. Porcine rotavirus passaged 10 times in cell culture infected gnotobiotic pigs and caused diarrhea. Gnotobiotic pigs that recovered from this infection were resistant to challenge exposure with porcine rotavirus but were susceptible to challenge exposure with transmissible gastroenteritis virus. As determined by immunofluorescent cross reactions, porcine rotavirus was found to be antigenically related to the human and bovine rotaviruses but not to reovirus type 3 or to transmissible gastroenteritis virus.     

间接免疫荧光方法快速检测番鸭呼肠孤病毒病

本研究应用具有免疫荧光特性的抗番鸭呼肠孤病毒病单克隆抗体(MDRV 237-11)建立了检测番鸭呼肠孤病毒病抗原的间接免疫荧光(Mab-IFA)方法。结果显示人工感染MDRVMW9710株病毒后发病和死亡番鸭组织切片均可检出阳性病灶;且组织匀浆经尿囊腔途径接种12日龄番鸭胚,取死亡胚心制作冰冻切片进行IFA鉴定呈阳性结果。表明该方法特异性好、可用于快速检测番鸭呼肠孤病毒病。     

Virologico-serologic studies in horses with respiratory tract diseases]

Of 1081 acute and chronically respiratory diseased as well as clinically normal horses 824 sera and 257 paired serum samples collected 1986 and 1987 were tested for antibodies against several different respiratory viruses such as influenza virus A/equi 1 and 2 (Influenza 1 a. 2), equine herpesvirus type 1/4 (EHV 1/4), mammalian reovirus type 1-3 (Reovirus 1-3), equine rhinovirus type 1 (ERV 1), equine adenovirus type 1 (EAdV 1), and equine arteritis virus (EAV). The investigations resulted in an antibody prevalence of 57.2% (Influenza 1), 59.5% (Influenza 2), 81.5% (EHV 1/4), 50.3% (Reovirus 1), 43.0% (Reovirus 2), 75.9% (Reovirus 3), 97.6% (EAdV 1), 82.5% (ERV 1) and 8.7% (EAV). With exception of EAV and EAdV 1 the ratios usually were higher in diseased animals than in clinically normal horses. Antibodies to EAV and EAdV 1 were present in all groups to almost the same amount. Of 257 horses with acute respiratory illness 3 showed a significant rise of the antibody titer against Influenza 1, 30 against Influenza 2, 54 against EHV 1/4, 1 against Reovirus 1 and 3, respectively, 11 against EAdV 1 and 26 against ERV 1.     

Cell Culture Studies of a Neonatal Calf Diarrhea Virus

The effects of a neonatal calf diarrhea virus on cell cultures were investigated. Bovine embryonic kidney cell cultures were the most satisfactory for production of virus. Cytoplasmic changes detected after inoculation with a high multiplicity of virus were: 1) cytoplasmic vacuoles; 2) some eosinophilic cytoplasmic inclusions; and 3) some degeneration of cells and detachment from the monolayer. Cultures stained with fluorescein-labeled antibody showed cytoplasmic fluorescence as early as four hr after infection with the maximum fluorescence at five days. No cross reactions were observed between the neonatal calf diarrhea virus and reovirus type 1 or type 3 by the fluorescent antibody technique. Plaques were small and were not produced consistently. The optimal adsorption time was one to two hr. The maximum titer was reached at 18 hr, with the cell-associated titer remaining higher than the cell-free titer until that time. An interferon was produced by cultures infected with either ultraviolet-inactivated or untreated virus.     

Effect of a live reovirus vaccine on reproductive performance of broiler breeder hens and development of viral tenosynovitis in progeny.

A live commercial reovirus vaccine, Enterovax, was administered to adult broiler breeder hens via the drinking water to determine its efficacy in stimulation of circulating antibody. This vaccine was compared with a commercial inactivated reovirus vaccine. Only the inactivated product resulted in increased antibody as measured by a commercial enzyme-linked immunosorbent assay. However, the live reovirus vaccine caused diarrhea in the hens and decreased eggshell quality, fertility, and hatchability. In addition, the live vaccine virus was vertically transmitted from hens to their progeny, resulting in increased embryonic mortality and viral tenosynovitis.