Interferon Production and Host Resistance to Type II Avian (Marek's) Leukosis Virus (JM Strain)

Interferon production in both susceptible S- and resistant K-line chickens infected with type II leukosis virus of JM strain and turkey herpesvirus was studied. The resistant line of chickens produced higher levels of interferon than did the susceptible with JM virus infection during the experimental period. When both susceptible S-and resistant K-line chicks were vaccinated with turkey herpesvirus, the interferon production was quantitatively similar in the two lines.     

Comparison of Indirect Hemagglutination and Immunodiffusion Tests for Detecting Type II Leukosis (Marek's) Infection in S- and K-Line Chickens

The indirect hemagglutination and immunodiffusion tests were compared for detection of antigen and antibody to JM strain of leukosis virus infection between S- and K-line chickens. The indirect hemagglutination test was more sensitive than the immunodiffusion test for detecting the smallest amount of viral antigen and corresponding antibody in the plasma of infected chickens. The Cornell S-line had higher levels of antigen and antibody as compared with the Cornell K-line during the 20-week experimental period.     

Comparison of Indirect Hemagglutination and Immunodiffusion Tests for Detecting Type II Leukosis (Marek''s) Infection in S- and K-Line Chickens

The indirect hemagglutination and immunodiffusion tests were compared for detection of antigen and antibody to JM strain of leukosis virus infection between S- and K-line chickens. The indirect hemagglutination test was more sensitive than the immunodiffusion test for detecting the smallest amount of viral antigen and corresponding antibody in the plasma of infected chickens. The Cornell S-line had higher levels of antigen and antibody as compared with the Cornell K-line during the 20-week experimental period.     

Lyophilization of Cell-Free Marek''s Disease Herpesvirus and a Herpesvirus from Turkeys

Cell extracts of the JM and GA strains of Marek's disease herpesvirus and the FC 126 strain of turkey herpesvirus were lyophilized with various stabilizers. Much higher virus titers were obtained with stabilizer than without stabilizer. Titers increased even further in the case of the Marek's disease virus strains by the addition of a chelating agent, disodium ethylenediaminetetraacetate.     

Marek's disease virus and herpesvirus of turkey noninfective to chickens, obtained by repeated in vitro passages.

The relation between the passage level of Marek's disease virus, C2 strain, and of herpesvirus of turkey (HVT), O1 strain, in cell culture and the level of the serological response of chickens to these viruses was examined. In both cases the immune response of chickens to these viruses decreased with increase in the number of in vitro passages of virus. Virus was not recovered from chickens inoculated with HVT highly passaged in vitro, which had become a high producer of cell-free virus in vitro, and grew equally well at 37 C and 41 C.     

Lyophilization of Cell-Free Marek's Disease Herpesvirus and a Herpesvirus from Turkeys

Cell extracts of the JM and GA strains of Marek''s disease herpesvirus and the FC 126 strain of turkey herpesvirus were lyophilized with various stabilizers. Much higher virus titers were obtained with stabilizer than without stabilizer. Titers increased even further in the case of the Marek''s disease virus strains by the addition of a chelating agent, disodium ethylenediaminetetraacetate.     

Protective Effect of Avian Myelomonocytic Growth Factor in Infection with Marek’s Disease Virus

Marek's disease virus (MDV) is a herpesvirus that induces T lymphomas in chickens. The aim of this study was to assess the role of the macrophage activator chicken myelomonocytic growth factor (cMGF) in controlling MDV infection. B13/B13 chickens, which are highly susceptible to MD, were either treated with cMGF delivered via a live fowlpox virus (fp/cMGF) or treated with the parent vector (fp/M3) or were left as untreated controls. Seven days later, when challenged with the very virulent RB-1B strain of MDV, the spleens of chickens treated with fp/cMGF showed increased expression of the inducible nitric oxide synthase (iNOS) gene compared to those of control chickens and fp/M3-treated chickens. Increased iNOS gene expression was also accompanied by greater induction of gamma interferon and macrophage inflammatory protein (K203) gene expression, both possible activators of iNOS. fp/cMGF treatment also increased the number of monocytes and systemic NO production in contrast to fp/M3 treatment. Even though cMGF treatment was unable to prevent death for the chickens, it did prolong their survival time, and viremia and tumor incidence were greatly reduced. In addition, cMGF treatment improved the partial protection induced by vaccination with HVT (herpesvirus isolated from turkeys) against RB-1B, preventing 100% mortality (versus 66% with vaccination alone) and greatly reducing tumor development. Treatment with fp/M3 did not have such effects. These results suggest that cMGF may play multiple roles in protection against MD. First, it may enhance the innate immune response by increasing the number and activity of monocytes and macrophages, resulting in increased NO production. Second, it may enhance the acquired immune response, indicated by its ability to enhance vaccine efficacy.     

Augmentation of retrovirus-induced lymphoid leukosis by Marek''s disease herpesviruses in White Leghorn chickens.

Our objective was to determine whether the cell-associated herpesvirus vaccines used in chickens to control Marek's disease tumors can augment development of lymphoid leukosis (LL) induced by exogenous avian leukosis virus (ALV). Various single or mixed Marek's disease vaccines were inoculated at day 1, and ALV was injected at 1 to 10 days, with chickens of several experimental or commercial strains. Development of LL was monitored at 16 to 48 weeks in various experiments. In several strains of chickens we repeatedly found that the widely used serotype 3 turkey herpesvirus vaccine did not augment LL in comparison with unvaccinated controls. However, LL development and incidence were prominently augmented in several chicken strains vaccinated with serotype 2 vaccines, used alone or as mixtures with other serotypes. In one chicken strain, augmentation was demonstrated after natural exposure to ALV or serotype 2 Marek's disease virus viremic shedder chickens. Augmentation of LL by virulent or attenuated Marek's disease viruses of serotype 1 was intermediate in effect. Serotype 2 Marek's disease virus augmentation of LL was prominent in three laboratory lines and one commercial strain of White Leghorns, but it was not observed in an LL-resistant laboratory line or four commercial strains susceptible to ALV infection. Chickens developed similar levels of viremia and neutralizing antibodies to ALV regardless of the presence of augmentation of LL, suggesting that the mechanism of enhanced LL did not result from differences in susceptibility or immune response to ALV. We postulate that the serotype 2 herpesviruses may augment LL through one of several possible influences on bursal cells that are subsequently transformed by exogenous ALV.     

火鸡疱疹病毒在鸡胚成纤维细胞上有效代次的测定及其免疫原性恢复的研究

本试验证明,火鸡疱疹病毒(HVT)FC126株在鸡胚成纤维细胞(CEF)传55代的病毒,仍有预防马立克氏病(MD)的效力,至第70代效力明显下降。若将HVT CEF第35代病毒复归火鸡连续传4代,能明显恢复其免疫原性,用C细胞传代则有效代次可达到75代。因而,此方法能有效地解决免疫原性下降的种毒的更新问题。本试验还为该种毒和疫苗的有效代次的使用范围提供了可靠依据。     

Avian leukosis virus infection: analysis of viremia and DNA integration in susceptible and resistant chicken lines

Avian leukosis viruses induce lymphoid leukosis, a lymphoma which develops within the bursa of Fabricius several months after virus infection. Chickens from the Hyline SC and FP lines are, respectively, susceptible and resistant to avian leukosis virus-induced lymphoid leukosis. We examined plasma and cellular DNA obtained from avian leukosis virus-infected chickens for the presence of viremia and integrated viral sequences to determine whether the extent of virus infection is comparable in individuals of both lines. A less than twofold difference in the frequency of viremia was detected between chickens of the two different lines. Although the analysis of plasma samples, which were obtained at different times postinfection, demonstrated that the duration of viremia was comparable in both susceptible and resistant chickens, the onset of the viremia observed in susceptible chickens generally preceded by 1 week that observed in resistant chickens. Moreover, integrated viral sequences were detected in approximately 90% of the SC and 40% of the FP chickens. The appearance of infectious virus in the plasma was, in general, associated with the presence of integrated viral sequences in both the bursal cells and the erythrocytes obtained from the same chicken. The presence of both the viremia and the integrated viral DNA sequences was transient, suggesting a mechanism for the elimination of virus-infected cells in both susceptible and resistant chickens. Furthermore, at 5 weeks postinfection no integrated exogenous viral sequences were detected in splenic lymphocytes obtained from either chicken line, regardless of whether these chickens were viremic or had integrated viral sequences detectable in other tissues. Our results indicate that extensive avian leukosis virus replication occurs in approximately 50% of the FP and 100% of the SC chickens. Although it appears that the viral infection spreads more quickly in the SC chickens, our results afford no obvious explanation of the resistance to the development of lymphoma exhibited by FP chickens.     

Interferon and Host Resistance to Rauscher Virus-induced Leukemia

A random bred strain of mice (CD-1) was shown to develop resistance to Rauscher leukemia virus (RLV) as the animals matured. Resistant adult mice developed relatively high-serum levels of interferon (150 to 2,000 units per ml) in contrast to susceptible 21-day-old animals in which interferon levels were undetectable or low (less than 20 to 200 units per ml). A similar correlation between resistance and interferon levels was observed in comparisons between resistant CD-1 and susceptible BALB/c mice. The F(1) hybrids of CD-1 x BALB/c and BALB/c x CD-1 matings manifested an intermediate degree of susceptibility and interferon production. The difference in interferon production by CD-1 and BALB/c mice was specific for the RLV-host interaction, since both strains produced equal serum levels of interferon in response to Sindbis and Newcastle disease viruses. The mortality of CD-1 suckling mice infected with Rauscher leukemia virus was decreased by treatment with interferon. These data demonstrate an association between interferon production by the host and the observed relative resistance of the CD-1 strain of adult mice to the subsequent malignant transformation. This virus-host relationship provides an excellent model for further study of factors affecting the development of virus-induced leukemia.     

8-Azaguanine resistant African green monkey kidney cell mutants (Vero 153): isolation and characterization

A clone of Vero cells resistant to up to 20 micrograms/ml 8-azaguanine was isolated. This clone (designated Vero 153) has a doubling rate of approximately 24 h and a maximum cell density of 10,000/mm2. Deficiency of the enzyme hypoxanthine guanine phosphoribosyl transferase (HGPRT) in Vero 153 was demonstrated by methods of radiochromatography. Vero 153 is susceptible to hypoxanthine-thymidine-aminopterin (HAT) medium and its resistance to 8-azaguanine seems to be nonreversible. Like parental cells, Vero 153 was also incapable of interferon production when challenged with Newcastle disease virus (NDV) or poly(inosinic acid) . poly(cytidylic acid) (poly I:C). Similar chromosome complements (majority range 56 to 57) and band patterns were observed in cells harvested at Passages 10, 20, and 50. The potential use of Vero 153 for somatic cell hybridization for purposes of gene mapping, virus rescue, and the control of interferon production is discussed.     

Entamoeba histolytica: genetic control of susceptibility in chicken eggs.

Five strains of axenized Entamoeba histolytica were established in eggs from chickens of different genetic backgrounds. The eggs of outbred and inbred chickens varied in the proportion of embryos from which the strains of E. histolytica were recovered and there were variations in the numbers of trophozoites recovered from individual positive eggs. It was possible, however, to find individual, single-mated hens which laid only uniformly susceptible or resistant eggs. Susceptibility and resistance to E. histolytica could be demonstrated in embryonated eggs derived both from outbred heterogenous and in highly inbred chickens but eggs from a genetically stable source were relatively constant in the distribution of susceptible and resistant embryos. Except for gross hydrocephalus, tissue reactions were not observed in the chick embryos or in the chorioallantoic, amniotic, or yolk sac membranes. The yolk sac inoculation route and 37 C incubation temperature provided the optimal survival conditions for the amoebae. Infection with avian leukosis virus and interferon production did not seem to play a role in susceptibility and resistance. Neither the presence of E. histolytica cross-reacting, maternal antibody from the hen transmitted transovarially via the yolk sac, or other antiamoebic substances appeared to play a role in the resistance observed in nonsusceptible eggs. Multiple alternating passages of trophozoites in hen's eggs/TPS-1 broth led to an increase in the number of positive embryos and a decrease in embryo deaths.     

Non-association between Rfp-Y major histocompatibility complex-like genes and susceptibility to Marek's disease virus-induced tumours in 63× 72 F2 intercross chickens

Marek's disease (MD) is a lymphoproliferative disease caused by a member of the herpesvirus family, and the best understood genetic resistance to MD involves the chicken major histocompatibility complex (MHC) B -complex. Preliminary observations have suggested that MHC-like Rfp-Y genes might also influence the incidence of MD. This study describes the differentiation and definition of unique Rfp-Y genes in inbred lines 6₃ and 7₂, lines that possess identical B -complex genes, but that are resistant or susceptible to MD, respectively. To assess if Rfp-Y genes affect susceptibility to MD, 265 6₃× 7₂ F₂ chickens were challenged with the JM strain of MD virus at 1 week of age and were evaluated for MD lesions at up to 10 weeks of age. Genotyping of the F₂ chickens for Rfp-Y haplotypes was performed by restriction fragment length polymorphism analysis of genomic DNA using Taq I and a B-FIV probe. Analysis of variance and interval mapping procedures were used to determine association between the Rfp-Y haplotypes and the phenotypic MD values of the F₂ chickens. The cosegregation analysis of 265 F₂ chickens indicated that there was no association between Rfp-Y haplotypes and MD susceptibility. Furthermore, the fact that the Rfp-Y haplotypes fit the 1:2:1 segregation ratio and the Rfp-Y allele frequencies did not differ significantly from 0·5 in the full population or in selected subpopulations (of either 40 MD-resistant or 39 MD-susceptible chickens) also indicated that Rfp-Y haplotypes do not significantly influence MD susceptibility. We conclude that Rfp-Y haplotypes do not play a major role in determining the genetic susceptibility to MD in 6₃× 7₂ F₂ White Leghorn chickens.     

The effect of substrate on the production of infectious virus by cells in culture

Herpes simplex virus type I (HSV-1), infectious bovine rhinotracheitis virus (IBR) and turkey herpesvirus were examined for growth in cells cultured on three different substrates. The substrates were glass, DEAE-dextran and collagen gel. With two of the viruses, HSV-1 and IBR, there were no apparent differences in production as a function of substrate. In contrast, the amount of the turkey herpesvirus which was recovered varied greatly with the substrate. Titers were highest on glass, followed by DEAE-dextran and then collagen gel. Our previous studies have indicated that the substrate on which anchorage-dependent cells are grown in vitro has an affect on a number of biological and biochemical properties. The present study indicates that the production of commercially important biologicals can be affected by the substrate.     

Inhibitory effects of nitric oxide and gamma interferon on in vitro and in vivo replication of Marek's disease virus

The replication of Marek's disease herpesvirus (MDV) and herpesvirus of turkeys (HVT) in chicken embryo fibroblast (CEF) cultures was inhibited by the addition of S-nitroso-N-acetylpenicillamine, a nitric oxide (NO)-generating compound, in a dose-dependent manner. Treatment of CEF culture, prepared from 11-day-old embryos, with recombinant chicken gamma interferon (rChIFN-gamma) and lipopolysaccharide (LPS) resulted in production of NO which was suppressed by the addition of N(G)-monomethyl L-arginine (NMMA), an inhibitor of inducible NO synthase (iNOS). Incubation of CEF cultures for 72 h prior to treatment with rChIFN-gamma plus LPS was required for optimal NO production. Significant differences in NO production were observed in CEF derived from MDV-resistant N2a (major histocompatibility complex [MHC], B(21)B(21)) and MDV-susceptible S(13) (MHC, B(13)B(13)) and P2a (MHC, B(19)B(19)) chickens. N2a-derived CEF produced NO earlier and at higher levels than CEF from the other two lines. The lowest production of NO was detected in P2a-derived CEF. NO production in chicken splenocyte cultures followed a similar pattern, with the highest levels of NO produced in cultures from N2a chickens and the lowest levels produced in cultures from P2a chickens. Replication of MDV and HVT was significantly inhibited in CEF cultures treated with rChIFN-gamma plus LPS and producing NO. The addition of NMMA to CEF treated with rChIFN-gamma plus LPS reduced the inhibition. MDV infection of chickens treated with S-methylisothiourea, an inhibitor of iNOS, resulted in increased virus load compared to nontreated chickens. These results suggest that NO may play an important role in control of MDV replication in vivo.     

Characteristics of resistance in rice to rice hoja blanca virus (RHBV) and its vector, Tagosodes orizicolus (Muir)

This paper characterises resistance to rice hoja blanca virus (RHBV) which is transmitted by the planthopper Tagosodes orizicolus (Muir). Resistance is expressed as decreased proportion of plants infected compared to susceptible lines, although severity of symptom expression is similar in both types. This resistance is not due to differences in vector feeding behaviour. Vectors reared eight generations on resistant plants showed no increased ability to transmit to resistant lines or decreased ability to transmit to susceptible ones. Longevity of vectors was similar when reared either on virus-resistant or susceptible plants. Incubation period of the virus in resistant plants are significantly longer than in susceptible plants. Resistance increased with plant age in resistant and susceptible cultivars. Increased virus dosage, as determined by increased number of viru-liferous vectors per inoculated plant, caused an increase in transmission to both resistant and susceptible cultivars. However, the ranking of resistant and susceptible remained the same across the experimental range of dosage and plant age. It is concluded that the resistance studied is to virus infection and there is little risk of “breakdown” occurring as a result of genetic or behavioural changes in the vector population. This will permit the use of economic thresholds to planthopper feeding damage with little risk of RHBV epidemic outbreak.     

Map location of homologous regions between Marek''s disease virus and herpesvirus of turkey and the absence of detectable homology in the putative tumor-inducing gene.

The DNA region having homology between Marek's disease virus and herpesvirus of turkey was assigned to the restriction map of Marek's disease virus by Southern blot hybridization. Under moderate conditions at the level of 15% mismatching, homology was found to be distributed throughout the Marek's disease virus genome. The long inverted-repeat regions (TRL and IRL), which are considered to play a significant role in tumorigenicity, did not show any homology to herpesvirus of turkey DNA.