Systemic Spread Of Virus Research Articles

Resistance to Systemic Spread of High Plains Virus and Wheat Streak Mosaic Virus Cosegregates in Two F2 Maize Populations Inoculated with Both Pathogens

High plains virus (HPV) has the potential to cause significant damage in susceptible maize (Zea mays L.) genotypes. The virus is vectored by the wheat curl mite (WCM), Aceria tosichella, Keifer, which also vectors wheat streak mosaic virus (WSMV). We have previously characterized susceptibility of maize inbred lines to double infection of HPV and WSMV. The objective of this study was to characterize the inheritance of resistance to systemic spread of HPV and WSMV in mixed infection. Genetic analysis was done with crosses B73 (resistant) × Wf9 (susceptible) an B73 × W64A (susceptible). Parental, F1, and F2 plants were WCM‐inoculated with HPV and WSMV, and scored for their reactions both visually and by enzymelinked immunosorbent assay. F1 plants were resistant to systemic spread of HPV and WSMV, indicating that resistance is dominant in these lines. Segregation (based on symptomology) in the F2 generation fit a 3:1 resistant:susceptible ratio in both B73 × W64A and B73 × Wf9 (P = 0.34 and 0.11, respectively), consistent with a single chromosome region segregating for resistance to both viruses. The resistant allele was contributed by B73 and was linked to marker bnl6.29 on the short arm of chromosome six. This chromosome region has been shown to control resistance to WSMV (wsm1). Two independent sets of inbred‐backcross‐derived near‐isogenic lines confirmed the effect of this chromosome region. This study reveals the genetic basis of symptomology observed upon mixed infection with HPV and WSMV in these populations. Identification of molecular markers linked to virus resistance genes may enhance inbred line development efforts.

Requirement of the Movement Protein for Long Distance Spread of Tobacco Mosaic Virus in Grafted Plants

Systemic spread of tobacco mosaic virus (TMV) that lacks a functional movement protein (TMVΔMP) was investigated in grafted tobacco (Nicotiana tabacum) plants. Transgenic plants that express the 30-kDa movement protein (MP) gene (MP) under the control of the rolC (phloem-specific) or pal2 (xylem-specific) promoters were unable to support systemic infection by the mutant virus, while plants that express the MP gene from the cauliflower mosaic virus 35S promoter (35S:MP) led to systemic infection. Doubly grafted plants were constructed in which plants containing the 35S:MP gene were used as root stock and plants carrying various MP constructs constituted the middle scion. The upper scion contained the 35S:MP gene in plants that produce a hypersensitive response when systemically infected by TMV. TMVΔMP moved systemically and produced complete necrosis in the upper scion when expression of MP in the middle scion was under the control of the rolC or 35S promoter, but not when the pal2 promoter was used. When plants expressing a gene encoding a defective MP were used as the middle scion, there was no systemic infection by TMVΔMP, and a delay in systemic infection by wild-type TMV. In grafted plants with middle scions that expressed the TMV 54 kDa gene sequence there was no apparent systemic infection by TMVΔMP in the upper scion. The results obtained indicate that the MP has a role in long distance movement, and support the suggestion that replication is necessary for systemic infection of these grafted plants.

Glycoprotein E2 of bovine viral diarrhea virus expressed in insect cells provides calves limited protection from systemic infection and disease.

Calves were vaccinated with a C-terminally truncated baculovirus expression product of E2 from the Singer strain of bovine viral diarrhea virus. The expressed E2 was glycosylated and retained antigenic authenticity. After induction of viral neutralizing antibody, the calves were challenge exposed with either the homologous Singer strain of virus or with the heterologous 890 strain of virus. Vaccine-induced antibody titer of > or = 2 protected calves from clinical signs of disease induced by homologous viral challenge exposure. An antibody titer of > or = 512 reduced replication of homologous challenge virus to a level which did not induce an appreciable increase in serologic titer of viral neutralizing antibody. Vaccine-induced antibody titer of < or = 4096 did not protect calves from systemic spread of virus or from disease after challenge exposure with heterologous bovine viral diarrhea virus.

An interspecies hybrid RNA virus is significantly more virulent than either parental virus.

Cucumber mosaic cucumovirus (CMV) infects a very wide range of plant species (>1000 species). We recently demonstrated that a previously undescribed gene (2b) encoded by RNA 2 of the tripartite RNA genome of CMV is required for systemic virus spread and disease induction in its hosts. Herein we report that when this CMV gene is replaced by its homologue from tomato aspermy cucumovirus (TAV), the resultant hybrid virus is significantly more virulent, induces earlier onset of systemic symptoms, and accumulates to a higher level in seven host species from three families than either of the parents. Our results indicate that CMV and the TAV 2b protein interact synergistically despite the fact that no synergism occurs in double infections with the two parental viruses. To our knowledge, this is the first example of an interspecific hybrid made from plant or animal RNA viruses that is more efficient in systemic infection of a number of hosts than the naturally occurring parents. As CMV and the hybrid virus accumulated to a similar level in the infected tobacco protoplasts, the observed synergistic responses most likely resulted from an increased efficacy of the hybrid virus in systemic spread in host plants provided by the TAV 2b protein. The relevance of our finding to the application of pathogen-derived resistance is discussed.

Movement of tomato mosaic virus strains inGomphrena globosaas related to temperature sensitivity and plant resistance

Symptom development, virus multiplication, and distribution of tomato mosaic virus strains L and LS1were compared at 22 and 32 °C in the C4plant Gomphrena globosa. In inoculated leaves, LS1accumulation was much lower at 32 °C compared with 22 °C, whereas the levels of L were similar at both temperatures. At 22 °C, spread of L and LS1from inoculated to uninoculated leaves (systemic movement) was dependent on the stage of plant development and the leaf inoculated. Under all conditions tested, LS1accumulated at a much lower level than L. Under specific conditions of leaf and plant age, LS1was confined to the inoculated leaf, whereas L moved systemically in all experiments. In a mixed infection with tobacco mosaic virus U1strain, LS1did not affect movement of U1within the inoculated leaf but inhibited systemic spread. In Gomphrena, the single amino acid difference in the 30-kDa movement protein of LS1relative to L induced plant resistance to systemic spread of virus. Keywords: systemic movement, local lesion.

Complementation of African cassava mosaic virus AC2 gene function in a mixed bipartite geminivirus infection.

We have previously demonstrated that African cassava mosaic virus (ACMV) DNAs A and B efficiently complement the systemic spread of tomato golden mosaic virus (TGMV) DNA A when co-agroinoculated onto Nicotiana benthamiana. Here, we show that a mixture of an ACMV DNA A AC2 mutant and DNA B that is normally unable to systemically infect N. benthamiana can do so at low frequency when co-agroinoculated with TGMV DNA A. Analysis of viral DNA showed that the AC2 mutation was retained during infection. The mixture of genomic components was sap transmissible, indicating that systemic infectivity is not specifically attributable to the use of agroinoculation. In the presence of TGMV DNA A, ACMV coat protein as well as the DNA B gene products BV1 and BC1 were detected in systemically infected tissues. The results demonstrate that dysfunctional AC2 can be complemented in planta by its TGMV homologue AL2.

Assessment of protection from systemic infection or disease afforded by low to intermediate titers of passively acquired neutralizing antibody against bovine viral diarrhea virus in calves

SUMMARY Colostrum-deprived calves (n = 24) were fed various amounts of colostrum, colostrum substitute, or milk replacer to establish a range in titer of passively acquired viral neutralizing antibody in serum. The calves were then challenge exposed intranasally with a virulent, noncytopathic bovine viral diarrhea virus (bvdv-890). After viral challenge exposure, calves were monitored for fever, leukopenia, thrombocytopenia, and diarrhea. In addition, viral isolation and viral titration were performed on specimens of nasal secretions, buffy coat cells, and serum obtained from the calves. Fever and systemic spread of virus were detected in calves that had viral neutralizing titer of 256 or lower. Calves that had viral neutralizing titer lower than 16 developed severe clinical disease manifested by fever, leukopenia, thrombocytopenia, and diarrhea. Severity and duration of signs of disease decreased as titers of passively acquired viral neutralizing antibody increased. These results indicate that low to intermediate titers of passively acquired viral neutralizing antibody were not sufficient to fully protect calves from virulent bovine viral diarrhea virus.

Mutational analysis of the movement protein of odontoglossum ringspot virus to identify a host-range determinant.

Tobacco mosaic virus (TMV) which contains the movement protein (MP) of odontoglossum ringspot tobamovirus (ORSV) in place of the TMV MP systemically infects orchids but causes local infection in tobacco unless the carboxy-terminal 48 amino acids of the MP are deleted (C. A. Holt, C. A. Fenczik, S. J. Casper, and R. N. Beachy; Virology, in press, 1995). Frameshift mutations were created within the 3' ends of the MP gene that led to truncations of the ORSV MP by 11, 19, 28, 37, and 48 amino acids; each of the mutant MP genes was inserted into the cloned cDNA of TMV in place of the TMV MP and infectious transcripts were produced. Virus containing mutant MPs were used to infect vanilla orchids, a systemic host of ORSV, and tobacco plants. Removal of 11 amino acids from the ORSV MP prevented spread of the chimeric virus in orchids while restoring the ability to cause a systemic infection on tobacco. Further deletions of the MP affected the size of virus-induced necrotic local lesions on tobacco cv. Xanthi NN and the systemic spread and accumulation of virus in cv. Xanthi nn, a systemic host of TMV. However, each virus replicated to equivalent levels in protoplasts. A mechanism by which the ORSV MP limits the spread of the chimeric virus is proposed.

Coding density of the turnip yellow mosaic virus genome: Roles of the overlapping coat protein and p206-readthrough coding regions

More than one-third of the turnip yellow mosaic virus (TYMV) genome simultaneously encodes two ORFs. We haveinvestigated the functions of the overlapping coat protein ORF and readthrough domain of ORF-206 in the 3′ region of the genome. TYMC-206 RNA, in which a second stop codon has been positioned to prevent ORF-206 readthrough, induced infections in protoplasts and plants that were indistinguishable from wild type. ORF-206 readthrough is thus nonessential. Nevertheless, TYMV-221 RNA, in which the ORF-206 stop codon was replaced with a tyrosine codon to force readthrough, was infectious to protoplasts, suggesting that a role for ORF-206 readthrough under certain conditions is possible. TYMV RNA variants that produce truncated or no coat protein were used to show that the coat protein is dispensable for local movement but necessary for systemic spread of virus in plants. Studies in protoplasts showed that (−) RNA levels are normal in the absence of coat protein, but (+) strand levels are decreased about 10-fold relative to wild-type infections. A mutant with a short C-terminal coat protein extension that formed virions less stable than normal demonstrated the protective role of capsids toward genomic RNA. The evolutionary implications of the dense information content of the TYMV genome are discussed.

Ultrastructure of Tobacco Ringspot Virus‐Induced Local Lesions in Lima Bean Leaves

AbstractTobacco ringspot virus (TRSV), a nepovirus, induced unusually large, expanding lesions in the primary leaves of lima bean (Phaseolus lunatus cv. Nemagreen). During the course of lesion development, two phases were identified on the basis of ultrastructural observations. In the primary phase, cytological changes followed a pattern that was characteristic of compatible or systemic nepovirus infections. Such changes included the formation of large, membranous inclusion bodies in the cytoplasm of mesophyll parenchyma and the accumulation of phytoferritin in the stroma of chloroplasts. The cells maintained their structural integrity and appeared metabolically active. The secondary phase was degradative in nature. Ultrastructural changes observed during this phase were representative of those that occur in a virus‐induced incompatible or hypersensitive (necrotic) reaction. Such changes included rounding of chloroplasts, vesiculation and disintegration of the plasma membrane and tonoplast, degeneration of cell organelles, and generalized plasmolysis. Subsequently, the cell walls collapsed, developing folds and loops around the shrunken cytosol. The collective result of these two ultrastructurally distinctive phases was the production of large necrotic lesions in the inoculated leaf, but no systemic viral spread.

The mode of cauliflower mosaic virus propagation in the plant allows rapid amplification of viable mutant strains.

We inoculated the leaves of turnip plants (Brassica campestris spp. rapa cv. Just Right) with two cauliflower mosaic viruses (CaMVs) with different small mutations in a dispensable region of the viral genome, and followed the spread of the virus infection through the plant. Surprisingly, analysis of viral DNA in single primary chlorotic lesions revealed the presence of both mutants. In contrast, the secondary chlorotic lesions and systemically infected leaves contained virus molecules of either one or the other type only. Infection of plants with different ratios of the two reporter viruses showed that this ratio is not conserved during systemic virus spread. Infection with CaMV DNA in the form of heteroduplexes containing a single mismatched base pair, in which each strand carried a distinct diagnostic marker, provided us with evidence that the mismatch was subjected to a repair process in the host plant.

Local and systemic spread of tobacco mosaic virus in transgenic tobacco.

Expression of a chimeric gene encoding the coat protein (CP) of tobacco mosaic virus (TMV) in transgenic tobacco plants confers resistance to infection by TMV. We investigated the spread of TMV within the inoculated leaf and throughout the plant following inoculation. Plants that expressed the CP gene [CP(+)] and those that did not [CP(-)] accumulated equivalent amounts of virus in the inoculated leaves after inoculation with TMV-RNA, but the CP(+) plants showed a delay in the development of systemic symptoms and reduced virus accumulation in the upper leaves. Tissue printing experiments demonstrated that if TMV infection became systemic, spread of virus occurred in the CP(+) plants essentially as it occurred in the CP(-) plants although at a reduced rate. Through a series of grafting experiments, we showed that stem tissue with a leaf attached taken from CP(+) plants prevented the systemic spread of virus. Stem tissue without a leaf had no effect on TMV spread. All of these findings indicate that protection against systemic spread in CP(+) plants is caused by one or more mechanisms that, in correlation with the protection against initial infection upon inoculation, result in a phenotype of resistance to TMV.

Local and Systemic Spread of Tobacco Mosaic Virus in Transgenic Tobacco

Local and Systemic Spread of Tobacco Mosaic Virus in Transgenic Tobacco

Protection against lethal influenza with neuraminidase

The role of the neuraminidase in eliciting protection against a lethal influenza A virus [A/Ck/Penn/1370/83 (H5N2)] infection was investigated in chickens. Isolated N2 neuraminidase administered in adjuvant did not prevent infection but did prevent systemic spread of virus and death of chickens. N2 expressed in a recombinant vaccinia virus protected chickens when administered in adjuvant but was less effective when allowed to replicate and produce pox on the chicken's comb. Chickens vaccinated with isolated N2 in adjuvant or with inactivated H5N2 influenza virus were protected from clinical signs and death after challenge with A/Ck/Penn/1370/83 influenza virus. However, these animals were completely susceptible and died of infection with a heterologous subtype (H7N7) of influenza virus. The role of the different antigenic determinants on the N2 NA was investigated in chickens by passive transfer of monoclonal antibodies. Antibodies to antigenic determinants rimming the enzyme active center reduced disease signs in approximately half of the birds but did not significantly reduce virus levels. Antibodies to one of the two independent antigenic determinants that are distant from the enzyme active center were most effective at reducing virus replication and disease signs. This is surprising because antigenic variants could not be selected in vitro with these antibodies and suggests that they may facilitate clearance of virus. Antibodies to the other determinant that is located distally to the enzyme site were ineffective at providing protection.

Central nervous system involvement in HIV infection.

Central nervous system (CNS) involvement occurs frequently in patients with the acquired immunodeficiency syndrome (AIDS), but at present only a few reports have addressed the analysis of intrathecal IgG synthesis in human immunodeficiency virus (HIV)-seropositive patients with no signs of HIV-related neurologic syndromes. In this study, intrathecal IgG synthesis was investigated using several techniques in patients with different stages of HIV infection and then correlated with the state of the blood-brain barrier. Almost all patients had specific anti-HIV IgG synthesis within the CNS, suggesting the presence of HIV in the brain. These findings further stress that direct CNS infection occurs early in the course of systemic virus spread.

Multiple components of the resistance of potatoes to potato leafroll virus

SUMMARYIn glasshouse experiments the ranking of potato genotypes for resistance to infection with potato leafroll virus (PLRV) using three concentrations of aphid‐borne inoculum was the same as their field resistance ratings. In field‐grown plants this resistance to infection increased in all genotypes as the plants aged but its rate of increase differed between genotypes. In tests on field‐grown plants infected by aphid‐ or graft‐inoculation, the proportion of virus‐free progeny tubers increased the later the date of inoculation but was greater in resistant than in susceptible genotypes. This trend was most pronounced in the resistant clone G7445(1), in which the virus failed to move from the foliage to the tubers of some plants infected in glasshouse tests.The spread of PLRV will thus be minimised in crops of resistant compared with susceptible genotypes for three reasons: plants have greater resistance to infection, systemic spread of virus from their foliage to tubers is less likely and, as shown previously, the low concentration of virus particles in leaf tissue makes infected plants less potent sources of inoculum for aphids.

Protection of guinea pigs against local and systemic foot-and-mouth disease after administration of synthetic lipid amine (Avridine) liposomes

Injection of the synthetic lipid amine, Avridine, in the form of liposomes, protected guinea pigs against the development of lesions from foot-and-mouth disease virus (FMDV) inoculated intradermally into the rear footpads. The animals were protected against the development of vesicles at the inoculation site as well as the systemic spread of virus. Maximal protection was obtained after intracardial injection of 5–10 mg doses of liposomal Avridine. Lower doses yielded decreased protection. Subcutaneous or intraperitoneal routes of liposomal Avridine injection were ineffective. Protection was maximal 0–24 h after injection of liposomes. Ethanol and emulsion formulations of Avridine could induce protection when injected intracardially but had toxic side effects. Guinea pigs protected against the first FMDV inoculation by liposomal and ethanol formulations of Avridine continued to be protected against lesions after a second inoculation 15–45 days later. FMDV protective antibody titers of these animals ranged from a low of < 1:10 to > 1:1000.

Protective role of foot-and-mouth disease virus antibody in vitro and in vivo in guinea-pigs.

SUMMARY Plaque reduction neutralization assays, using foot-and-mouth disease virus (FMDV) type A, strain 119 and immune serum from convalescent guinea-pigs infected with this strain of virus and performed with monolayers of a swine kidney cell line, resulted in biphasic neutralization curves because of the presence of as many as 30 to 50% of non-neutralized virus particles at peak activity. These results were found using gum tragacanth, agar, agar containing DEAE-dextran, and methylcellulose overlays and were also found using monolayers of guinea-pig embryo tongue and guinea-pig embryo heelpad cells. Non-neutralized virus in immune serum-FMDV mixtures was neutralized after the addition of anti-species antibody, demonstrating that the non-neutralized virus fraction consisted of virus in the form of infectious immune complexes. These complexes were not infectious when inoculated intraperitoneally into suckling mice or intracardially into guinea-pigs. They were infectious, however, if inoculated intradermally into the tongue or rear heelpads of guinea-pigs. Low doses of passively transferred immune serum did not protect guinea-pigs against the formation of primary vesicles after intradermal tongue or heelpad challenge with virus but did protect against systemic spread of virus to the remaining uninoculated feet. Higher doses of passively transferred immune serum protected against tongue challenge but even higher doses were required to protect against heelpad challenge. The role of antibody in protection against the systemic spread of FMDV may be due to infectious immune complexes being removed from the blood by the reticuloendothelial system. In the dermis of the tongue and heelpad, the immune complexes remain infectious, resulting in the formation of local vesicles except when these tissues contain very high concentrations of antibody.

Mechanisms underlying systemic invasion of pea plants by pea enation mosaic virus.

Leaves of Pisum sativum L.cv. 'Perfected Wales' were inoculated with pea enation mosaic virus, at the 3rd and 4th nodes as early as size permitted. Systemic virus spread was then measured over a period of approximately 9 days by assaying the infectivity of 1-cm stem pieces above the 4th and below the 3rd node. The presence of virus antigen and double-stranded (ds) RNA in these stem pieces was measured by radioimmunoassay over the same period. Relative infectivity and amounts of dsRNA in infected tissue were lowest approximately 3 days after inoculation and sharply increased thereafter. A transient plasmodesmatal abnormality was observed 4-6 days after inoculation. The transient nature of the abnormality, which was preceded (24 h) by low infectivity and dsRNA concentrations, may indicate a plant defense reaction that is subsequently overcome. The rise in dsRNA concentration, starting the 4th day after inoculation, coincided with the increase of vesicular material (cytopathological structures) in the phloem of infected plants. Vesicular material was found in sieve elements that were mature at the time of inoculation. Their presence in this tissue can only be explained by transport of the vesicles from infected sites.

Chemotherapy and plant viruses.

SUMMARY: When the guanine analogue, 5-amino-7-hydroxy-1-v-triazolo (d) pyrimidine (guanazolo), was sprayed on the leaves of tobacco or Nicotiana glutinosa plants it reduced the number of local lesions and delayed or inhibited systemic spread of lucerne mosaic virus. Guanazolo was more effective when applied before inoculation, but had some effect if applied up to about the second day after inoculation. The compound was more effective in solution in 0.1% sodium bicarbonate than in aqueous suspension. With mechanically inoculated plants guanazolo watered on the soil around the plants was less effective than when sprayed on the leaves. Incubated with the virus in vitro the compound did not affect infectivity. In concentrations up to c. 0.005m guanazolo usually caused negligible plant damage, but at higher concentrations produced a slight yellowing and distortion in the younger leaves with general stunting if treatments were prolonged. The virus-inhibitory activity of guanazolo was reversed by adenine, guanine and possibly by hypoxanthine, but not by xanthine, uric acid, theobromine, theophylline, caffeine, uracil or thymine. The triazolo analogue of adenine severely damaged plants and had only slight virus inhibitory activity. The hypoxanthine analogue caused no plant damage. It was less effective than guanazolo in tobacco and N. glutinosa but more effective in reducing the number of local lesions produced in beans. Thiouracil, methyl-thiouracil and propylthiouracil were ineffective against lucerne mosaic virus. Thiouracil caused fairly severe plant damage. Guanazolo had slight or negligible effects on spotted wilt virus in tomato, potato viruses X and Y in potato, and tobacco and pea mosaic virus in peas. Applied as a leaf spray at 0.01m concentration guanazolo delayed or prevented systemic movement of cucumber mosaic virus from mechanically inoculated cucumber leaves, but had no effect when watered on the soil. However, watering the compound on the soil gave some control when the virus was introduced by aphids.