Howard A. Scott

Degradation of matrix protein from a nuclear-polyhedrosis virus of Trichoplusia ni by an endogenous protease

Abstract The intact matrix protein of a nuclear-polyhedrosis virus of Trichoplusia ni could be isolated when a contaminating endogenous protease was inhibited by Hg 2+ . The subunit molecular weight of the intact matrix protein was approximately 28,000. When the protease was not inhibited, the matrix protein was degraded to a mixture of peptides which had a strong tendency to self-associate. The protease-degraded matrix protein preparation could be dissociated by 1.0 M formic acid into a single peptide and a peptide aggregate. The sum of the amino acid compositions of the peptide and peptide aggregate was identical, within experimental error, with the amino acid composition of the intact matrix protein.

A newly isolated densonucleosis virus from Pseudoplusia includens (Lepidoptera: Noctuidae)

Abstract An icosahedral DNA virus isolated from the soybean looper, Pseudoplusia includens , was characterized. Purified virus had a diameter of 20 ± 1 nm and negatively stained preparations showed a trend to form linear to three-dimensional crystals. The virus had a sedimentation coefficient of 120 ± 3 S and a buoyant density of 1.40 ± 0.01 g/cm 3 . The DNA content of the virus was 37.8 ± 0.1% and the absorption spectrum showed it to be a typical nucleoprotein. Viral DNA in situ was shown to be single-stranded by staining the virus with acridine orange as well as by reaction to formaldehyde. Evidence of inverted terminal repetition of the DNA was observed by electron microscopy. The terminal repetition comprises ca. 6–7% of the genome. The molecular weight of the ssDNA was 2.0 ± 0.1 × 10 6 as determined by agarose gel electrophoresis or 2.1 ± 0.1 × 10 6 as determined by electron microscopy. Four virion proteins with molecular weights of 46.5 ± 0.1, 54.0 ± 0.1, 64.0 ± 0.2, and 87.0 ± 0.1 × 10 3 were detected by 9% SDS-polyacrylamide gel electrophoresis. Double-diffusion tests showed the virus to be serologically related but not identical to DNV-1. Ultrathin sections showed that the nucleus of the hemocyte, muscle, hypodermal, and fat body cells contained virus-like particles. The chromatin of an infected nucleus always underwent a margination and the nucleoplasm was often replaced largely by virions. Data indicate that the virus belongs to the Densovirus of the family Parvoviridae.

Isolation and some properties of components of nuclear polyhedra from the cabbage looper, Trichoplusia ni

Abstract Nuclear polyhedra obtained from diseased cabbage looper, Trichoplusia ni, were digested with sodium carbonate-saline buffer, pH 11.0. The dissolved polyhedra formed 3 general zones when subjected to density gradient centrifugation. The slowest sedimenting component (Zone 1) had an ultraviolet absorption curve typical of protein and a sedimentation coefficient of 11 S. Capsids, 310 × 40 nm, were located in Zone 2. Virus particles were found in 1–3 bands (Zone 3); those with envelopes measured 300 × 72 nm, and those without envelopes measured 300 × 33 nm. Virus preparations stained with phosphotungstic acid at pH 7.0 exhibited extensive disruption whereas preparations stained at pH 3.0 did not. Virus particles in the sodium carbonate-saline-digested polyhedra had a sedimentation coefficient of 1228 S. Virus particles isolated by high speed centrifugation had a sedimentation coefficient of 1530 S.

Evidence that Ribonuclease in Beetle Regurgitant Determines the Transmission of Plant Viruses

SummarynRegurgitant from leaf-feeding beetles (Cerotoma trifurcata, Epilachna varivestis and Diabrotica undecimpunctata) contains RNase activity equivalent to 0.1 to 1 mg/ml pancreatic RNase. When mixtures of pancreatic RNase at 0.1 to 1.0 mg/ml and viruses that are or are not transmissible by beetles, were inoculated to plants using the gross wounding technique, those containing viruses not transmissible by beetles were not infective. Pancreatic RNase at activities up to 25 times that found in beetle regurgitant did not prevent transmission of two beetle-transmissible viruses, whereas RNase at activities one-half that found in beetle regurgitant prevented or greatly inhibited transmission of two viruses not transmitted by beetles.

Evidence that Virus Translocation and Virus Infection of Non-wounded Cells Are Associated with Transmissibility by Leaf-feeding Beetles

SummarynTwo experimental systems were used to study the translocation of virus particles in plants and translocation followed by infection of non-wounded plant cells. Translocation of virus particles in cut plant stems occurred with the beetle-transmissible viruses southern bean mosaic, bean pod mottle and cowpea severe mosaic, but not with sunn hemp mosaic and tobacco ringspot viruses, which are not transmitted by beetles. When purified viruses were injected below sections of steam-killed stems, only the beetle-transmissible viruses were translocated and infected non-wounded tissue above the steam-killed area. However, sunn hemp mosaic, tobacco ringspot and bean yellow mosaic viruses, which are not transmitted by beetles, did not infect plants above killed portions of the stem. These results suggest that both translocation of virus particles in the xylem of plants and establishment of infection in non-wounded tissue are key properties of viruses that are transmitted by beetles.

Comparison of the relationships of southern bean mosaic virus and the cowpea strain of tobacco mosaic virus with the bean leaf beetle.

Abstract Bean leaf beetles, Cerotoma trifurcata , transmitted southern bean mosaic virus (SBMV) but not the cowpea strain of tobacco mosaic virus (CP-TMV) when fed on infected tissue or injected with purified virus. Both viruses were found in hemolymph and induced regurgitant obtained from fed or injected beetles, and both were recovered from leaf surfaces in the areas where beetles had fed. Apparently, despite extensive feeding activities and the presence of virus in the vicinity of damaged tissues, SBMV but not CP-TMV was deposited at the proper site for infection to occur.

Osmiophilic Globules and Myelinic Bodies in Cells Infected with Two Comoviruses

SummarynBean leaf cells infected with bean pod mottle virus or cowpea mosaic virus contain osmiophilic globules (OG) in the cytoplasm. OG and myelinic bodies (MB) occurred between the plasmalemma and cell wall, and also embedded in the cell wall. Tubules containing virus particles were associated with OG and MB outside the cytoplasm, and membranes of some tubules were associated with MB. It is suggested that OG move from the cytoplasm through the plasmalemma, remain near the cell wall and become transformed into MB.

Immunoelectrophoresis of hemolymph of the cabbage looper, Trichoplusia ni, during the course of a nuclear polyhedrosis virus infection.

Larvae of the cabbage looper, Trichoplusia ni, were infected with a nuclear polyhedrosis virus and the protein patterns of the hemolymph followed throughout the course of the disease by immunoelectrophoresis. There was a decrease in the protein concentration of the hemolymph which appeared to be due to the loss of two normal protein components and to a reduction in the concentration of other major normal proteins. Proteins were detected following virus infection which were absent from hemolymph of healthy larvae. These proteins were not serologically related to those present in the virus.

Development of monoclonal antibodies specific for Pyricularia grisea, the rice blast pathogen

The development of polyclonal antisera and monoclonal antibodies (MAbs) specific for Pyricularia grisea was explored. Polyclonal antisera and MAbs against germinating conidia cross-reacted with unrelated fungi and extracts of healthy rice tissue in ELISA. However, MAbs produced from crushed conidia showed various levels of specificity to the fungus. Fourteen hybridoma lines secreting antibodies positive for the immunogen and negative for healthy rice tissue were selected from three independent fusions of NS-1 myeloma cells with splenocytes from mice immunized with crushed conidial suspensions of P. grisea race IB-49. Four stable cell lines were recloned. MAbs secreted from cell lines 4G11, 8H1 and 3E4 reacted strongly with conidial antigen, and MAb 11C6 preferably with mycelial antigen in ELISA. MAb 4G11 bound to the surface of conidia whereas 8H1 and 3E4 bound on germ-tubes in the indirect immunofluorescence assay (IFA). In cross-reaction tests with ELISA MAb 4G11, an IgG1 antibody, reacted negatively with isolates representing 11 fungal genera and was defined as species-specific. Further tests against 17 isolates of P. grisea indicated that MAb 4G11 reacted positively with 11 and 12 of the isolates in ELISA and IFA, respectively. MAb 4G11 could detect homologous conidial antigen at 14–70 ng ml −1 , 10–20 conidia/well, and the fungal antigen in infected rice tissue in ELISA. It may have potential diagnostic value.

Determinants in the Specificity of Virus Transmission by Leaf-feeding Beetles

Vector specificity is a well-recognized characteristic of plant viruses. The plant viruses that are transmitted by leaf-feeding beetles are in four virus groups and cannot be transmitted by other commonly recognized virus vectors such as aphids, leafhoppers, or nematodes. The basis for vector specificity has been elucidated for some virus-vector combinations, and in many cases the interaction between the vector and virus is determined by virus coat proteins or by nonstructural proteins, as with the aphid-transmitted potyviruses and caulimoviruses (16, 17). These viral proteins have been shown to mediate a specific, definitive interaction of the virus particle with some surface of the vector. Sites of virus binding to vector surfaces that play a role in vector specificity have been identified for nematode, fungal, aphid, and leafhopper vectors (2, 16, 17). In some cases, virus coat proteins are involved in interactions with and transport across vector membranes as in the case with luteovirus passage through the aphid gut wall and accessory salivary gland cells (15).