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More cell culture passaged Camelpox virus sequences found resembling those of vaccinia virus.

ABSTRACT: Background:Camelpox is the most infectious and economically important disease of camelids that causes significant morbidity and mortality rates. Several live attenuated vaccines against Camelpox virus (CMLV) are produced worldwide by passaging field isolates in cell culture. Sequence of a high passage Saudi isolate of CMLV was previously found closely resembled Vaccinia virus (VACV). Aim:To determine whether other high cell culture passage CMLV isolates are genetically resemble VACV and further to explore the possible mechanism of the resemblance. Methods:We performed polymerase chain reaction and DNA sequence analysis of A-type inclusion body protein (ATIP), L1R, and open reading frame (ORF) 185 genes on different cell culture passage levels of a field isolate, two high passage vaccines, wild-type, and reference strains of CMLV. Results:We demonstrate that additional two high passage attenuated vaccine candidate from Sudan and UAE likewise contain sequences resembling VACV more than CMLV. Furthermore, sequence analysis of the ATIP gene of selected virus passages in cell culture revealed that the shift to VACV-like occurred between passage 11 and 20 and up to the 10th passage the genome still resembles wild-type virus. This observation was further confirmed by recombination analysis which indicated recombination events at ATIP and ORF185 genes occurred at higher passages. Conclusion:We confirmed that the cell culture passage CMLV turns to resemble VACV after cell culture passage and concluded that the resemblance may not be a result of contamination or misidentification as previously thought but could be due to recombination events that occurred during the passage process.

SUBMITTER: Khalafalla AI

PROVIDER: S-EPMC7419068 | biostudies-literature | 2020 Aug

REPOSITORIES: biostudies-literature

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More cell culture passaged Camelpox virus sequences found resembling those of vaccinia virus.

Khalafalla Abdelmalik I AI Al Hosani Mohamed A MA Ishag Hassan Zackaria Ali HZA Al Muhairi Salama S SS

Open veterinary journal 20200423 2

<h4>Background</h4>Camelpox is the most infectious and economically important disease of camelids that causes significant morbidity and mortality rates. Several live attenuated vaccines against <i>Camelpox virus</i> (CMLV) are produced worldwide by passaging field isolates in cell culture. Sequence of a high passage Saudi isolate of CMLV was previously found closely resembled Vaccinia virus (VACV).<h4>Aim</h4>To determine whether other high cell culture passage CMLV isolates are genetically rese ...[more]

PMID: 32821659

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Project description:Kemerovo virus (KEMV) is a tick-borne orbivirus transmitted by ticks of the genus Ixodes. Previous animal experimentation studies with orbiviruses, in particular the interferon receptor double knock-out (IFNAR(-/-)) mouse model, did not indicate bias that is related to age or sex. We endeavoured to assess the effect of serial and alternated passages of KEMV in mammalian or Ixodes cells on virus replication and potential virulence in male or female IFNAR(-/-) mice, with important age differences: younger males (4-5 months old), older males (14-15 months old), and old females (14-15 months old). After 30 serial passages in mammalian or tick cells, or alternated passages in the two cell types, older female mice which were inoculated with the resulting virus strains were the first to show clinical signs and die. Younger males behaved differently from older males whether they were inoculated with the parental strain of KEMV or with any of the cell culture-passaged strains. The groups of male and female mice inoculated with the mammalian cell culture-adapted KEMV showed the lowest viraemia. While older female and younger male mice died by day 6 post-inoculation, surprisingly, the older males survived until the end of the experiment, which lasted 10 days. RNA extracted from blood and organs of the various mice was tested by probe-based KEMV real-time RT-PCR. Ct values of the RNA extracts were comparable between older females and younger males, while the values for older males were >5 Ct units higher for the various organs, indicating lower levels of replication. It is noteworthy that the hearts of the old males were the only organs that were negative for KEMV RNA. These results suggest, for the first time, an intriguing age- and sex-related bias for an orbivirus in this animal model. Changes in the amino acid sequence of the RNA-dependent RNA polymerase of Kemerovo virus, derived from the first serial passage in Ixodes cells (KEMV Ps.IRE1), were identified in the vicinity of the active polymerase site. This finding suggests that selection of a subpopulation of KEMV with better replication fitness in tick cells occurred.

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More cell culture passaged Camelpox virus sequences found resembling those of vaccinia virus.

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More cell culture passaged Camelpox virus sequences found resembling those of vaccinia virus.

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