Nonoccluded baculovirus- and filamentous virus-like particles in the spotted cucumber beetle,diabrotica undecimpunctata (coleoptera: chrysomelid)

Nonoccluded baculovirus-and filamentous virus-like particles were found in nuclei of hemocytes or midgut cells of field-collected spotted cucumber beetles. Each type of particle was associated with a different type of virogenic stroma containing various viral components similar to those referred to as capsid, nucleocapsid, viroplasm, and viral envelope in other known baculovirus infections. Nucleocapsids of the virus which occured only in hemocytes were rod-shaped particles approximately 230 nm long and 52 nm wide and were enveloped singly by a trilaminar unit membrane. Enveloped and partly enveloped particles appeared to be released from the nucleus to the cytoplasm by budding through the nuclear envelope acquiring additional membranes. The nucleocapsids of the virus which occurred only in nuclei of midgut cells were filamentous particles with an average diameter of 25 nm and variable length up to 2 μm. Some extremely long particles were bent almost 360° near the middle, resulting in a hairpin-like configuration. The particles were always enveloped singly. No particles budding through the nuclear envelope were observed.     

Histopathology of a nuclear polyhedrosis infection in Aedes epactius with observations in four additional mosquito species

Aedes epactius larvae were utilized to study the infection sequence of the nuclear polyhedrosis virus (NPV) from Aedes sollicitans. From 30 min to 6 hr postinoculation, polyhedra and many free virions were observed in the larval midgut lumen. Penetration of the midgut cells by virions was not observed. The first infected nuclei were observed 12 hr postinoculation. Nucleocapsids initially exhibited electron translucent cores which became electron dense before the nucleocapsids acquired an envelope. Envelope acquisition occurred through a process of de novo membrane morphogenesis. Occlusion of the singly embedded virions began by 18 hr postinoculation with the mature rough-surfaced polyhedra averaging approximately 1 by 2 μm. Unusually long nucleocapsids (approximately two or three times the length of other nucleocapsids) were only observed in late infection period nuclei. There was no evidence that long nucleocapsids represented an early developmental stage for nucleocapsids of standard length. Infection was restricted to midgut nuclei and gastric caecae cells. Infected early instar A. epactius larvae became moribund 36 to 40 hr postinoculation and infected midgut nuclei were observed to undergo lysis. The late stages of NPV infection were observed in larvae of A. annandalei, Wyeomyia smithii, Toxorhynchites brevipalpus, and Eretmapodites quinquevittatus. Virion development and occlusion in these species was basically identical to the sequence observed in A. epactius larvae.     

A nonoccluded virus of the army cutworm

A nonoccluded virus was isolated from larvae of the army cutworm, Euxoa auxiliaris. Infected larvae became lethargic and shrunken, and death usually occurred 12–20 days after infection. The primary site of viral infection and replication appeared to be the nuclei of midgut epithelial cells; however, virus replication also occurred in cells of the tracheal matrix and in muscle. Nuclei in early stages of the infection contained large granular areas with the chromatin scattered near the nuclear membrane. These areas differentiated into viral particles that measured 24 nm and formed crystalline arrays, occasionally 10 μm long. Disruption of the nuclear membrane liberated these arrays of particles into the cytoplasm. Fluorescence microscopy studies indicated that the viral particles contained DNA. The crystalline arrays were Feulgen positive. The virus also infected larvae of the armyworm, Pseudaletia unipuncta, and corn carworm, Heliothis zea, in laboratory tests.     

A new cytoplasmic polyhedrosis virus from the salt-marsh mosquito, Aedes cantator (Diptera: Culicidae)

The ultrastructure, development, and histopathology of a new cytoplasmic polyhedrosis virus of Aedes cantator are described. Virus particles measure 70 nm in diameter, are icosahedral in shape, and consist of a central electron-dense core surrounded by a capsid with six projections. Occlusion bodies are irregular in size (0.5–3.0 μm) and shape and contain several virus particles. Virus particles are assembled within an interconnecting network of fine filaments and are occluded by the deposition of a proteinaceous crystal around groups of mature virus particles within a virogenic stroma. Infections are confined to cells of the cardia, gastric ceca, and posterior portion of the midgut, which hypertrophy and frequently lyse. Infected larvae die during the fourth larval instar or as pupae. The prevalence of infection in natural field populations is less than 1%.     

A baculovirus of the European skipper, Thymelicus lineola (Lepidoptera: Hesperiidae)

The nuclear polyhedrosis virus (NPV) of the European skipper, Thymelicus lineola, observed for the first time in Quebec in 1974, is highly pathogenic for its host. The infected larvae fill with pure polyhedra and die within 4 to 10 days. All tissues were infected except nerve cells and silk glands, but all nuclei of infected cells were filled with polyhedra. Biochemical analyses revealed that important metabolic disturbances occurred in infected larvae, such as serious modifications in the activity of certain enzymes. Polyhedra measured from 350 to 1330 nm in diameter and contained up to about 80 single-enveloped virions which measured to 270 × 58 nm. Abnormally short and abnormally long particles were also observed. Safety tests on mammals, fish, and beneficial insects revealed that this virus had no effect on these organisms, thus, it was recommended for the control of T. lineola outbreaks.     

Herpesvirus salmonis: Characterization of a New Pathogen of Rainbow Trout

A new agent, provisionally designated Herpesvirus salmonis, was isolated from post-spawning rainbow trout (Salmo gairdneri) and studied primarily in the RTG-2 rainbow trout cell line. Infection of RTG-2 cells resulted in the formation of syncytia and Cowdry type A intranuclear inclusions. Replication occurred regularly at 5 and 10°C, but was inconsistent at 15°C, largely inhibited at 0°C, and completely inhibited at 20°C or higher. The virus was acid, heat, ether, and chloroform labile, but stable to freezing and thawing. It did not hemagglutinate. Viral DNA had a buoyant density of 1.709 g/cm³ and a guanine-cytosine value of 50%. Hexagonal nucleocapsids had a diameter of 90 nm and were first seen in nuclei at 36 h. Enveloped forms measured about 150 nm and occurred both cytoplasmically and extracellularly. At 10°C, a one-step growth culture required about 96 h; cell-associated virus peaked at about 10⁵ PFU/ml and exceeded released virus by a factor of about 10.     

Cytopathology of the soybean looper,Pseudoplusia includens,infected with the Pseudoplusia includens icosahedral virus

Ultrastructural responses of soybean looper cells of various tissues infected with Pseudoplusia includens icosahedral virus (PIIV), a newly characterized RNA virus [Y. C. Chao, H. A. Scott, and S. Y. Young (1983)J. Gen. Virol.64, 1835–1838], were studied in situ. Most cells of fat body and epidermis consistently contained virus particles and associated cytopathic structures. Virus particles, corresponding to those of purified PIIV in morphology and size, always occurred in the cytoplasm either in membrane-bound virogenic stroma and/or freely in the ground cytoplasm. Virogenic stroma, which appeared to be distinct from those induced by other insect viruses, consisted of electron-dense matrix material, in which virus particles were embedded, and membranous vesicles, 70 or 80 nm in diameter, containing nucleic acid-like fibrils. Virus particles in virogenic stroma occurred as hexagonally arranged crystalline arrays made up primarily of homogeneously dense particles, while those in the ground cytoplasm were dispersed randomly and had an electron-lucent central core. Extremely large numbers of virus particles were also located in noncellular cuticle layers of the integument.     

Morphogenesis of Respiratory Syncytial Virus in a Green Monkey Kidney Cell Line (Vero)

The structure and morphogenesis of respiratory syncytial (RS) virus particles in a green monkey kidney cell line (Vero) were examined. Infected cells contained dense intracytoplasmic inclusions composed of filamentous structures. In places where inclusion material was associated with membranes, structural modifications were induced. There was a thickening of the membrane and an addition of projections 12 to 15 nm in length. The same changes were most frequently observed after association of isolated filamentous structures with the cytoplasmic membrane. The budding-off process was clearly visualized. The diameter of mature virus particles varied between 90 and 130 nm and that of the internal component varied between 11 and 15 nm. The similarities between ultrastructural features of cells infected with RS virus and pneumonia virus of mice are pointed out. It is proposed that these two viruses should be classified together in a third subgroup of myxoviruses.     

A new virus disease in the housefly, Musca domestica (Diptera)

Reovirus particles were isolated from adults in laboratory colonies of the housefly, Musca domestica. These particles were spherical in outline, 57–76 nm in diameter, and were found only in hemocyte cytoplasm, where virions have been disclosed by a new technique. Virions were present in large numbers, and viral inclusion bodies were identified. The virus particles had pentagonal and hexagonal shapes resembling a simple icosahedral structure. The virus was shown to be infectious and pathogenic to adult flies through injection or by feeding them suspensions from flies that had died of the virus. Electron micrographs of midgut sections from infected flies showed that the midgut cells were packed with dark undulating threads which were not present in uninfected flies. However, no virus particles or inclusion bodies could be seen in these cells. On the basis of their association with infected flies, and the similarity to results from other studies on reoviruses and insect viruses, it is suggested that these threads are an alternative replicative form of the reovirus. When the virus suspensions from heavily infected flies were dialyzed against weak alkaline solutions, the threads showed an inner component of coiled material, 12 nm in diameter, inside an envelope with a diameter of 50–83 nm, mean 60.3 ± 7.5, composed of subunits 7–8 nm long and 7–8 nm across.     

An enzootic nuclear polyhedrosis virus of pink shrimp: Ultrastructure,prevalence, and enhancement

A nuclear polyhedrosis virus exists in pink shrimp, Penaeus duorarum, from waters of the northern Gulf of Mexico. This virus is rod-shaped, 269 nm long, and possesses an outer envelope surrounding its nucleocapsid. The nucleocapsid is 50 nm in diameter. The virus occurs in nuclei of host hepatopancreatic and midgut cells, and is both free in the nucleus and occluded within pyramidal-shaped polyhedral inclusion bodies (PIB's). Histochemically and ultrastructurally, the shrimp PIB's appear to be ribonucleoprotein and in fine structure bear close resemblance to polyhedral inclusion bodies of Baculovirus species from insects. However, the lattice line-to-line spacing is greater than that usually reported for insect PIB's. Crowding and chemical stress of shrimp in aquaria may enhance and increase the virus infection and prevalence. In limited experiments, shrimp fed heavily infected hepatopancreatic tissues had much higher mortality than controls fed only fish. The virus appears to be enzootic in pink shrimp in nature. Cytopathological changes in infected cells of shrimp appear similar to those in insects infected with certain species of Baculovirus. The name Baculovirus penaei n.sp. is proposed for the shrimp virus.     

Purification and properties of elderberry latent virus, one of four sap-transmissible viruses obtained from American elder (Sambucus canadensis L.)

Cherry leaf roll, tomato black ring and two previously unrecorded viruses were transmitted to Chenopodium quinoa from Sambucus canadensis plants imported from the U.S.A. Of the two newly recognized viruses one, code-named elderberry virus A, has filamentous particles about 650 times 15 nm; the other, named elderberry latent virus (ELV), was transmitted to several herbaceous species but remained symptomless in elder and most other hosts. In C. quinoa sap ELV lost infectivity after dilution to 10^--⁵ to 10^-- ⁶ , 10 min at 85–90°C, and 7 days at 18°C. Infectivity of nucleic acid extracts was abolished by ribonuclease in 0.2 m sodium chloride. ELV was purified from C. quinoa leaf extracts that were clarified with chloroform, by precipitation at pH 5 and differential centrifugation. Purified preparations contained numerous isometric particles c. 30 nm in diameter and a few particles c. 17 nm in diameter. In 0.06 M phosphate buffer ELV sedimented as a major 112 S (calculated for infinite dilution) component and a 48 S minor component. ELV showed no serological relationship to twenty-seven other isometric plant viruses. Its present cryptogram is R/I: *I*:S/S:S/*.     

Histopathology of virus-like particles in Heliothis spp

Larvae of Heliothis spp. collected from cotton, soybean, and peanut fields in South Carolina were found to be infected with virus-like particles (VLPs). Infected larvae became pale and swollen, stopped feeding, and remained alive for 2–3 weeks. Hemolymph from these larvae was milky and contained numerous spherical bodies ranging in diameter from 2 to 10 μm. The hemolymph also contained VLPs which were oval and measured 375 × 125 nm. Infectivity tests with crude saline extracts of infected larvae demonstrated that the pathogen could be transmitted by injection but not per os. The spherical bodies contained VLPs (387 × 149 nm) surrounded by two envelopes and packed together in clusters. These VLPs were also found in fat body cells, cuticular epithelial cells, tracheal cells, and connective tissue associated with the body wall and the gut. They were not found in muscle tissue or in midgut epithelial cells. Similar VLPs have been found in Heliothis zea from Mississippi and Trichoplusia ni from California, but a positive identification of the VLPs has not been made in any of these studies. Morphologically they appear to be distinct from any other previously described insect viruses.     

Ultrastructural identification of a herpes-like virus infection in common carp Cyprinus carpio in Korea

We report on a herpes-like virus, which was found to be associated with mass mortality of common carp Cyprinus carpio for the first time in Korea in 1998. The external signs of infection in moribund fish were darkened coloration and severe branchial necrosis in the gill. Transmission electron microscopy revealed the presence of herpes-like viruses in spleen tissue. Infected spleen cells showed hypertrophied nuclei and degeneration. Numerous nucleocapsids of about 82 nm in diameter were found within the nucleus and cytoplasm of infected cells, and extracellular enveloped particles were also observed. We conclude that this virus was a likely significant cause of the high mortality of common carp in Korea in 1998.     

Virus-like particles in phloem tissue of chervil (Anthriscus cerefolium) infected with carrot red leaf virus

Isometric virus-like particles c. 22–25 nm in diameter were found in ultrathin sections of chervil leaves infected with carrot red leaf virus (CRLV). The particles were confined to the phloem and occurred in less than 5% of the cells in the vascular bundles. They were commonest in companion cells, occurred frequently in sieve elements and were also found in phloem parenchyma cells. The observations support other evidence that CRLV should be classified in the luteovirus group.     

Genomics and proteomics of Apis mellifera filamentous virus isolated from honeybees in China

Apis mellifera filamentous virus (AmFV) is a large DNA virus that is endemic in honeybee colonies. The genome sequence of the AmFV Swiss isolate (AmFV CH–C05) has been reported, but so far very few molecular studies have been conducted on this virus. In this study, we isolated and purified AmFV (AmFV CN) from Chinese honeybee (Apis mellifera) colonies and elucidated its genomics and proteomics. Electron microscopy showed ovoid purified virions with dimensions of 300–500×210–285 nm, wrapping a 3165×40 nm filamentous nucleocapsid in three figure-eight loops. Unlike AmFV CH–C05, which was reported to have a circular genome, our data suggest that AmFV CN has a linear genome of approximately 493 kb. A total of 197 ORFs were identified, among which 36 putative genes including 18 baculoviral homologs were annotated. The overall nucleotide similarity between the CN and CH–C05 isolates was 96.9%. Several ORFs were newly annotated in AmFV CN, including homologs of per os infectivity factor 4 (PIF4) and a putative integrase. Phylogenomic analysis placed AmFVs on a separate branch within the newly proposed virus class Naldaviricetes. Proteomic analysis revealed 47 AmFV virion-associated proteins, of which 14 had over 50% sequence coverage, suggesting that they are likely to be main structural proteins. In addition, all six of the annotated PIFs (PIF-0–5) were identified by proteomics, suggesting that they may function as entry factors in AmFV infection. This study provides fundamental information regarding the molecular biology of AmFV.     

Auswirkungen des Anbaus einer Wirts- und einer Nichtwirtspflanze unter wechselnden ökologischen Bedingungen auf die Populationsdynamik von Rhizoctonia solani Kühn und Fusarium solani f. pisi (Jones) Snyd. et Hans

An isometric virus was isolated from Helianthus annuus L. plants showing a yellow leaf spot mosaic on affected leaves. Infected plants were found in different ecological regions of Ukraine. A procedure of virus purification is described. The diametres of the virus particles were nonuniform and ranged from 50 to 120 nm. The sedimentation coefficient of the virus was 518–540 S and the floating density in the CsCl gradient was 1.22 g/cm³. The MW of proteins separated by electrophoresis amounted to 78±0.9, 58±0.8, 52±0.2, and 27±0.8 kDa, respectively. The virus was assigned to the tospoviruses for which sunflower is a new previously undescribed natural host plant.     

Mycoplasma-like bodies in plants infected with potato witches' broom disease and the response of plants to tetracycline treatment

The virus origin of a Czechoslovak isolate of potato witches' broom disease is discounted: electron micrographs of ultrathin section of phloem tissues from plants infected with potato witches' broom disease demonstratedMycoplasma-like bodies, spherical or elongated showing binary fission and fragmentation. The minute corpuscles have a diameter of about 50–60 nm, the largest bodies of 1000 nm. The width of elongated filamentous structures is about 200 nm, most oval bodies have a diameter of 250 nm. A weak tetracycline treatment of diseased plants causes a delay of symptom development; a strong dose of tetracycline (applied by means of the wick method into the stem) inhibits symptom appearance completely. Tetracycline produces a phytotoxic effect inhibiting the growth of tomato plants and causing (at higher concentrations) necrosis and death of these plants. There is a note in the paper dealing with the term “mycoplasma”. The word mycoplasma in the sense ofEriksson (1897) or ofMereschkowsky (1910) does not correspond to the genus nameMycoplasma Nowak (1929).     

Mechanical Transmission, Purification and Properties of an Isolate of Maize Stripe Virus from Venezuela

A Venezuelan isolate of maize stripe virus (MStpV) was successfully transmitted mechanically and by the leafhopper Peregrinus maidis from field infected plants to sweet cv. Iochief. After purification of maize infected with MStpV, fine spiral filamentous particles about 4 nm in diameter and with variable lengths were consistently associated with a nucleoprotein band present in CsCl or Cs₂SO₄ isopycnic gradients. Purified preparations exhibited a typical nucleoprotein absorption spectrum with a maximum at 260–263 nm and a minimum at 240–243 nm and a 260–280 ratio of 1.38. The density of the nucleoprotein in CsCl gradients was estimated at 1.29 g/ml. The sedimentation coefficient was calculated at 62 S. The nucleoprotein consisted of 5 % single stranded RNA and a capsid protein of molecular weight 33.500 daltons. Large quantities of non-capsid proteins were isolated from infected tissue with a molecular weight of 17.500 and 16.500 daltons. Peregrinus maidis, injected with purified MStpV preparation failed to transmit the disease to healthy plants. However, they were infectious when injected with clarified infected plant sap. Antisera against capsid and non-capsid proteins from MStpV-Florida strain reacted positively with the Venezuelan antigens.     

Donut-shaped “miniparticles” in nuclei of human and rat cells

Donut-shaped “miniparticles” were extracted from nuclei of various types of human and rat cells. Electron-microscopic investigations showed these particles were predominantly in sucrose density gradient fractions that had an approximate sedimentation coefficient of 21S. These particles were 113±8_A^o in diameter and had an electron dense center of 29±6_A^o. They appeared to be composed of 8 subunits. Quantitative analysis of the number of these particles by electron-micrographic field counting showed nuclei of tumor samples had a larger amount of the particles than the cytosol. However, normal cell cytosol had a larger number of particles than the nuclei. A group of proteins in the 25, 000–33, 000 molecular weight range was shown to be the main protein component by two dimensional gel electrophoresis.