scholarly journals A novel Betaretrovirus discovered in cattle with neurological disease and encephalitis

Retrovirology ◽  
2021 ◽  
Vol 18 (1) ◽  
Author(s):  
Melanie M. Hierweger ◽  
Michel C. Koch ◽  
Ronja V. Kauer ◽  
Zoltán Bagó ◽  
Anna Oevermann ◽  
...  

Abstract Background The majority of emerging infectious diseases in humans are of animal origin, and many of them are caused by neuropathogenic viruses. Many cases of neurological disease and encephalitis in livestock remain etiologically unresolved, posing a constant threat to animal and human health. Thus, continuous extension of our knowledge of the repertoire of viruses prone to infect the central nervous system (CNS) is vital for pathogen monitoring and the early detection of emerging viruses. Using high-throughput sequencing (HTS) and bioinformatics, we discovered a new retrovirus, bovine retrovirus CH15 (BoRV CH15), in the CNS of a cow with non-suppurative encephalitis. Phylogenetic analysis revealed the affiliation of BoRV CH15 to the genus Betaretrovirus. Results BoRV CH15 genomes were identified prospectively and retrospectively by PCR, RT-PCR, and HTS, with targeting of viral RNA and proviral DNA, in six additional diseased cows investigated over a period of > 20 years and of different geographical origins. The virus was not found in brain samples from healthy slaughtered control animals (n = 130). We determined the full-length proviral genomes from six of the seven investigated animals and, using in situ hybridization, identified viral RNA in the cytoplasm of cells morphologically compatible with neurons in diseased brains. Conclusions Further screening of brain samples, virus isolation, and infection studies are needed to estimate the significance of these findings and the causative association of BoRV CH15 with neurological disease and encephalitis in cattle. However, with the full-length proviral sequences of BoRV CH15 genomes, we provide the basis for a molecular clone and further in vitro investigation. Graphical Abstract

2021 ◽  
Author(s):  
Melanie M. Hierweger ◽  
Michel C. Koch ◽  
Ronja V. Kauer ◽  
Zoltán Bagó ◽  
Anna Oevermann ◽  
...  

Abstract Background: The majority of emerging infectious diseases in humans are of animal origin, and many of them are caused by neuropathogenic viruses. Many cases of neurological disease and encephalitis in livestock remain etiologically unresolved, posing a constant threat to animal and human health. Thus, continuous extension of our knowledge of the repertoire of viruses prone to infect the central nervous system (CNS) is vital for pathogen monitoring and the early detection of emerging viruses. Using high-throughput sequencing (HTS) and bioinformatics, we discovered a new retrovirus, bovine retrovirus CH15 (BoRV CH15), in the CNS of a cow with non-suppurative encephalitis. Phylogenetic analysis revealed the affiliation of BoRV CH15 to the genus Betaretrovirus.Results: BoRV CH15 genomes were identified prospectively and retrospectively by PCR, RT-PCR, and HTS, with targeting of viral RNA and proviral DNA, in six additional diseased cows investigated over a period of >20 years and of different geographical origins. The virus was not found in brain samples from healthy slaughtered control animals (n=134). We determined the full-length proviral genomes from six of the seven investigated animals and, using in situ hybridization, identified viral RNA in the cytoplasm of cells morphologically compatible with neurons in diseased brains.Conclusions: Further screening of brain samples, virus isolation, and infection studies are needed to estimate the significance of these findings and the causative association of BoRV CH15 with neurological disease and encephalitis in cattle. However, with the full-length proviral sequences of BoRV CH15 genomes, we provide the basis for a molecular clone and further in vitro investigation.


1984 ◽  
Vol 3 (13) ◽  
pp. 3049-3053 ◽  
Author(s):  
P. Vos ◽  
J. Verver ◽  
P. van Wezenbeek ◽  
A. van Kammen ◽  
R. Goldbach

Viruses ◽  
2021 ◽  
Vol 13 (12) ◽  
pp. 2425
Author(s):  
Willian P. Paim ◽  
Mayara F. Maggioli ◽  
Shollie M. Falkenberg ◽  
Akhilesh Ramachandran ◽  
Matheus N. Weber ◽  
...  

Bovine serum has been widely used as a universal supplement in culture media and other applications, including the manufacture of biological products and the production of synthetic meat. Currently, commercial bovine serum is tested for possible viral contaminants following regional guidelines. Regulatory agencies’ established tests focused on detecting selected animal origin viruses and are based on virus isolation, immunofluorescence, and hemadsorption assays. However, these tests may fail to detect new or emerging viruses in biological products. High-throughput sequencing is a powerful option since no prior knowledge of the viral targets is required. In the present study, we evaluate the virome of seven commercial batches of bovine serum from Mexico (one batch), New Zealand (two batches), and the United States (four batches) using a specific preparation and enrichment method for pooled samples and sequencing using an Illumina platform. A variety of circular replicase-encoding single-stranded (CRESS) DNA families (Genomoviridae, Circoviridae, and Smacoviridae) was identified. Additionally, CrAssphage, a recently discovered group of bacteriophage correlated with fecal contamination, was identified in 85% of the tested batches. Furthermore, sequences representing viral families with single-stranded DNA (Parvoviridae), double-stranded DNA (Polyomaviridae and Adenoviridae), single-stranded RNA (Flaviviridae, Picornaviridae, and Retroviridae), and double-stranded RNA (Reoviridae) were identified. These results support that high-throughput sequencing associated with viral enrichment is a robust tool and should be considered an additional layer of safety when testing pooled biologicals to detect viral contaminants overlooked by the current testing protocols.


2020 ◽  
Author(s):  
Yanjie Zhao ◽  
Weijun Su ◽  
Shuai Li

AbstractHere, we developed two novel methods to construct high content nanobody library in mammalian cells. For the first method, we employed a linear-double-stranded DNA based AND-gate (LBAG) strategy. Upstream- and downstream-linear-double-stranded DNAs (ldsDNAs), containing identical overlapping sequences, were co-transfected into cultured cell lines to conduct AND-gate calculation to form intact nanobody expression cassette. For the second method, we generated full-length nanobody expression ldsDNA by in vitro ligation of restrict cut up- and down-stream ldsDNAs. Then the ldsDNAs were directly transfected into mammalian cells to express nanobody library. Both methods generated over a million different nanobody sequences as revealed by high-throughput sequencing (HTS).


2014 ◽  
Vol 95 (9) ◽  
pp. 2071-2074 ◽  
Author(s):  
Rodion V. Gorchakov ◽  
Robert B. Tesh ◽  
Scott C. Weaver ◽  
Farooq Nasar

The genus Negevirus consists of insect-only viruses isolated from mosquitoes and sandflies. Here, we report the successful construction of a full-length infectious cDNA clone of Negev virus (NEGV) strain M30957. Viral RNA was transcribed in vitro and virus was readily rescued with or without the use of a cap analogue. These results strongly suggest that NEGV, and likely other members within the genus, is a non-segmented, single-stranded, positive-sense RNA virus.


Virology ◽  
1979 ◽  
Vol 96 (1) ◽  
pp. 88-99 ◽  
Author(s):  
Sue Batt-Humphries ◽  
Christian Simonsen ◽  
Ellie Ehrenfeld

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Alberto A. Amarilla ◽  
Julian D. J. Sng ◽  
Rhys Parry ◽  
Joshua M. Deerain ◽  
James R. Potter ◽  
...  

AbstractThe current COVID-19 pandemic is caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). We demonstrate that despite the large size of the viral RNA genome (~30 kb), infectious full-length cDNA is readily assembled in vitro by a circular polymerase extension reaction (CPER) methodology without the need for technically demanding intermediate steps. Overlapping cDNA fragments are generated from viral RNA and assembled together with a linker fragment containing CMV promoter into a circular full-length viral cDNA in a single reaction. Transfection of the circular cDNA into mammalian cells results in the recovery of infectious SARS-CoV-2 virus that exhibits properties comparable to the parental virus in vitro and in vivo. CPER is also used to generate insect-specific Casuarina virus with ~20 kb genome and the human pathogens Ross River virus (Alphavirus) and Norovirus (Calicivirus), with the latter from a clinical sample. Additionally, reporter and mutant viruses are generated and employed to study virus replication and virus-receptor interactions.


Author(s):  
C. Jennermann ◽  
S. A. Kliewer ◽  
D. C. Morris

Peroxisome proliferator-activated receptor gamma (PPARg) is a member of the nuclear hormone receptor superfamily and has been shown in vitro to regulate genes involved in lipid metabolism and adipocyte differentiation. By Northern analysis, we and other researchers have shown that expression of this receptor predominates in adipose tissue in adult mice, and appears first in whole-embryo mRNA at 13.5 days postconception. In situ hybridization was used to find out in which developing tissues PPARg is specifically expressed.Digoxigenin-labeled riboprobes were generated using the Genius™ 4 RNA Labeling Kit from Boehringer Mannheim. Full length PPAR gamma, obtained by PCR from mouse liver cDNA, was inserted into pBluescript SK and used as template for the transcription reaction. Probes of average size 200 base pairs were made by partial alkaline hydrolysis of the full length transcripts. The in situ hybridization assays were performed as described previously with some modifications. Frozen sections (10 μm thick) of day 18 mouse embryos were cut, fixed with 4% paraformaldehyde and acetylated with 0.25% acetic anhydride in 1.0M triethanolamine buffer. The sections were incubated for 2 hours at room temperature in pre-hybridization buffer, and were then hybridized with a probe concentration of 200μg per ml at 70° C, overnight in a humidified chamber. Following stringent washes in SSC buffers, the immunological detection steps were performed at room temperature. The alkaline phosphatase labeled, anti-digoxigenin antibody and detection buffers were purchased from Boehringer Mannheim. The sections were treated with a blocking buffer for one hour and incubated with antibody solution at a 1:5000 dilution for 2 hours, both at room temperature. Colored precipitate was formed by exposure to the alkaline phosphatase substrate nitrobluetetrazoliumchloride/ bromo-chloroindlylphosphate.


2008 ◽  
Vol 35 (S 01) ◽  
Author(s):  
H Leske ◽  
A Baiker ◽  
C Schichor ◽  
J.C Tonn ◽  
R Goldbrunner ◽  
...  

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