Vacuoles Of Infected Cells Research Articles

Morphological analysis for two types of viral particles in vacuoles of SARS-CoV-2-infected cells.

In this study, we analyzed the morphological structure of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in human cells. We identified the two types of viral particles present within the vacuoles of infected cells. Using transmission electron microscopy, we observed that SARS-CoV-2 particles exhibited both low- and high-electron-density structures, which was further confirmed through three-dimensional reconstruction using electron tomography. The budding of these particles was exclusively observed within these vacuoles. Intriguingly, viral particles with low-electron-density structures were confined to vacuoles, whereas those with high-electron-density structures were found in vacuoles and on the cell membrane surface of infected cells. Notably, high-electron-density particles found within vacuoles exhibited the same morphology as those outside the infected cells. This observation suggests that the two types of viral particles identified in this study had different maturation status. Our findings provide valuable insights into the molecular details of SARS-CoV-2 particles, contributing to our understanding of the virus.

Quantification of the Volume and Surface Area of Symbiosomes and Vacuoles of Infected Cells in Root Nodules of Medicago truncatula

Legumes are able to form endosymbiotic interactions with nitrogen-fixing rhizobia. Endosymbiosis takes shape in formation of a symbiotic organ, the root nodule. Medicago truncatula (M. truncatula) nodules contain several zones representing subsequent stages of development. The apical part of the nodule consists of the meristem and the infection zone. At this site, bacteria are released into the host cell from infection threads. Upon release, bacteria are surrounded by a host cell–derived membrane to form symbiosomes. After release, rhizobia grow, divide, and gradually colonize the entire host cell of the fixation zone of root nodules. Therefore, mature infected cells contain thousands of symbiosomes, which remain as individual units among other organelles. Visualization of the organization and dynamics of the symbiosomes as well as other organelles in infected cells of nodules is essential to understand mechanisms regulating the development of endosymbiosis between plants and rhizobia. To examine this highly dynamic developmental process we designed a useful imaging technique that is based on confocal scanning microscopy combined with different fluorescent dyes and GFP-tagged proteins (Gavrin et al., 2014). Here, we describe a protocol for microscopic observation, 3D rendering, and volume/area measurements of symbiosomes and other organelles in infected cells of M. truncatula root nodules. This protocol can be applied for monitoring the development of different host-microbe interactions whether symbiotic or pathogenic.

Molecular studies on Tomato aspermy virus isolates infecting chrysanthemums

Tomato aspermy virus (TAV), an important viral pathogen of chrysanthemums, was detected by double antibody sandwich-enzyme linked immunosorbent assay (DAS-ELISA) from chrysanthemums exhibiting mottling and deformed inflorescence grown in various states of India. Out of 15 cultivars (cvs.) tested, 11 (73.3%) were found to be positive for TAV. On sap inoculation the virus produced local lesions on Chenopodium spp. and Cucumis sativus. Nicotiana clevelandii, N. glutinosa, N. megalosiphon and N. tabacum reacted systemically to the virus producing severe mosaic, leaf deformation and characteristic leaf enations. Tomato plants produced malformed fruits (2-3/plant) with a few seeds. Myzus persicae and Aphis gossypii transmitted the virus non-persistently. Electron microscopy of partially purified viral preparations revealed polyhedral virions (ca. 29 nm dia). Cytopathology of infected leaves of N. clevelandii showed crystalline inclusions containing virions in the central vacuole of infected cells. The virions showed very good clumping with antiserum to TAV in liquid phase immuno-electron microscopy. Slot blot hybridization was performed to detect the virus in various chrysanthemum cultivars. The virus positivity was confirmed by reverse transcription-polymerase chain reaction (RT-PCR) and nucleotide sequencing of coat protein gene (CP). The amplified 657 bp fragment was about 97% identical with respect to CP sequences of other TAV isolates available in the database. In terms of derived amino acid sequence similarity, the homology value was 99%. In order to amplify RNA 1, RNA 2 and RNA 3, multiplex RT-PCR was performed. High genetic similarities of CP gene of TAV Indian isolates with that of other isolates of TAV indicate their probable common ancestry.

Description of Tropheryma whipplei gen. nov., sp. nov., the Whipple's disease bacillus.

A detailed characterization was performed of the Whipple's disease bacillus, strain Twist-MarseilleT, isolated from the cardiac valve of a patient with Whipple's disease bacillus endocarditis. This strain was isolated and maintained on human embryonic lung fibroblast monolayers, but could not be cultivated in the absence of living eukaryotic cells. Two morphological forms were observed, with differing staining properties; an intracellular form with intact and degenerating bacteria within vacuoles of infected cells and an extracellular form with masses of bacteria embedded in an extracellular matrix. Determination of the DNA G+C content confirmed that it belongs to the high-G+C gram-positive bacteria. Strain Twist-MarseilleT (= CNCM I-2202T) is proposed as the type strain of a new species within a new genus, Tropheryma whipplei gen. nov., sp. nov., that was provisionally created solely on the basis of 16S rRNA gene sequence data.

Properties and Cytopathology of a Tymovirus Isolated from Eggplant

An isometric virus measuring 28 nm was isolated from eggplant (Solanum melongena) and mechanically transmitted to several species of the families Amaranthaceae, Chenopodiaceae, Labiatae, and Solanaceae. The virus was vectored by the flea beetle, Epitrix fuscula. Ultrastructural studies showed that chloroplasts of infected plants were clustered and formed tymovirus-characteristic double memebrane-bound vesicles. In Nicotiana rustica, immunogold labeling studies showed that the largest numbers of virus particles were in the cytoplasm and vacuole of infected cells (...)

Isolation of Carnation Ringspot Virus from a Canal Near a Sewage Plant: cDNA Hybridization Analysis, Serology and Cytopathology

AbstractA small water sample of only 60 ml from the Oker Aue canal near Braunschweig was ultracentrifuged. The resuspended pellet was rubbed on Chenopodium quinoa leaves which responded with the formation of almost 200 local lesions. The virus causing these lesions was identified as carnation ringspot virus (CarRSV) by means of dot blot hybridization using random‐primed cDNAs to the viral nucleic acids and by means of serology. Northern blot analysis revealed that the two RNA species of the virus which consisted of c. 3.7 and 1.5 Kb, respectively, have little or no base sequence homology. In immunoelectrophoresis at pH 7.0 the virus migrated towards the cathode. The isometric particles were 33 nm in diameter, round to slightly angular in outline, and showed a distinct granular or knobly surface structure. Virus particles occurred in the cytoplasm, nuclei, and vacuoles of infected cells which in addition contained amorphous granules in the cytoplasm and/or proliferated endoplasmic reticulum. Heavily, affected cells were necrotized and contained large virus particle aggregates which sometimes were crystallized.CarRSV is the third carnation virus, after carnation mottle and carnation Italian ringspot viruses, which was identified in a natural water.

Chlamydia psittaci: inclusion vacuole ultrastructure.

The inclusion ultrastructure of fibroblasts infected with Chlamydia psittaci (6BC) was studied. Electron microscopic techniques were used which permitted the observation of whole infected host cells and 1.0-micron sectioned preparations. It was shown that the cytoplasmic inclusion vacuoles of infected cells contained interconnecting structures within which chlamydiae reproduce.

Unstable infectivity and sedimentable ds-RNA associated with lettuce speckles mottle virus

Infectivity associated with extracts of lettuce speckles mottle virus (LSMV)-infected tissues is unstable and is susceptible to short periods of aging in vitro or low concentrations of RNase. Infected leaves extracted using phenol yielded greater infectivity than comparable tissue extracted with buffer. No typical virus particles were observed in infected tissues or clarified preparations, but 50- to 70-nm spherical membranous particles were observed associated with the tonoplast in vacuoles of infected cells. Polyacrylamide gel electrophoresis of nucleic acids from infected tissue and LSMV-clarified preparations revealed abundant double-stranded RNA (ds-RNA) in both preparations. A species of LSMV-specific single-stranded RNA (ss-RNA) was also present in LSMV bentonite-clarified preparations. Infectivity was associated only with the ss-RNA fraction. Quantitative isolation of ds-RNA from whole tissue and clarified preparations showed the majority of the ds-RNA was recovered from the bentonite-clarified fractions, and this RNA appeared to be associated with a relatively small structure of a low buoyant density. These data suggest that LSMV lacks a functional coat protein and the sedimentable dsRNA fraction from LSMV-infected tissues more likely contains a viral-specific replication site and not true virus particles.