Development of real-time PCR assay for specific detection of cowpox virus

Gavrilova, Elena V.; Щербаков, Д. Н.; Maksyutov, Rinat А.; Щелкунов, С. Н.

doi:10.1016/j.jcv.2010.06.003

Cited by 17 publications

(13 citation statements)

References 40 publications

(69 reference statements)

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“…For many years, OPV infections have been diagnosed by serological and biological methods in many countries. Nowadays, molecular diagnostic techniques, especially real-time PCR, are becoming more popular as they are fast and reliable, allowing this approach to be successfully applied to ORV and CPXV [19]. Orthopoxvirus infection can be identified using the conserved viral growth factor gene as the target for PCR.…”

Section: Discussionmentioning

confidence: 99%

Skin lesions caused by Orthopoxvirus in children

Mazur‐Melewska

Pieczonka-Ruszkowska

Szpura

et al. 2020

pdia

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Introduction: The global eradication of smallpox and abandonment of mandatory smallpox vaccination has led to an increased proportion of the population who are immunologically naïve to infections caused by Orthopoxviruses (OPV). Aim: To present the different courses of OPV infection in children and to highlight the diagnostic difficulties in their differentiation from the other inflammatory processes. Material and methods: We retrospectively evaluated the medical documentation of 5 children with OPV infection. Clinical diagnosis of OPV infection was based on evaluation of animal contact and skin symptoms, characterised by either a single ulcer or disseminated lesions. In all five cases, blood samples and skin swabs were collected from the lesion(s) to identify specific OPV DNA fragments (Vgf, b9R and D11L genes) using PCR. Results: Two children presented with high fever, a single ulcer on the skin and local lymphadenopathy. The three other patients were in good general health and their skin lesions presented as a disseminated vesicular rash. Using the Vgf gene as the target for PCR, OPV infection was confirmed in material collected from skin lesions of all children and in blood samples of 4 children. The B9R and d11L genes tested positive in the skin material of 2 children and blood samples of 2 children. All analysed patients presented a history of ineffective antibiotic therapy. Conclusions: In the case of unclear necrotising skin lesions in children, the primary diagnosis always includes bacterial dermatitis. However, if the patient has come into contact with animals, diagnosis of OPV infection should also be considered.

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Section: Discussionmentioning

confidence: 99%

Skin lesions caused by Orthopoxvirus in children

Mazur‐Melewska

Pieczonka-Ruszkowska

Szpura

et al. 2020

pdia

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“…DNA was extracted from multiple specimens by using the QIAGEN QIAamp DNA Mini Kit. The DNA was analyzed by OPV-specific real-time PCR (rpo 18) [ 13 ], CPXV-specific real-time PCR [ 14 ], and real-time PCR detecting the cellular c-myc gene for relative quantification [ 15 ]. Additionally, conventional PCR was performed, amplifying the open reading frame (ORF) of the hemagglutinin (HA) gene followed by sanger sequencing [ 15 ].…”

Section: Methodsmentioning

confidence: 99%

Seasonal recurrence of cowpox virus outbreaks in captive cheetahs (Acinonyx jubatus)

Stagegaard¹,

Kurth

Stern

et al. 2017

PLoS ONE

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Cowpox virus infections in captive cheetahs (Acinonyx jubatus) with high morbidity and mortality have already been reported in the UK and Russia in the 1970s. However, most of the reported cases have been singular events. Here, we report a total of five cowpox virus outbreaks in cheetahs in the same safari park in Denmark between 2010 and 2014. Nine cheetahs showed varying severity of clinical disease; two of them died (22%). All episodes occurred between August and October of the respective year. No other carnivores kept at the same institution nor the keepers taking care of the animals were clinically affected. The clinical picture of cowpox was confirmed by extensive laboratory investigations including histopathological and molecular analyses as well as cell culture isolation of a cowpox virus. High anti-orthopoxvirus antibody titers were detected in all 9 diseased cheetahs compared to seven contact cheetahs without clinical signs and 13 cheetahs not in direct contact. Additionally, whole genome sequencing from one sample of each cluster with subsequent phylogenetic analysis showed that the viruses from different outbreaks have individual sequences but clearly form a clade distinct from other cowpox viruses. However, the intra-clade distances are still larger than those usually observed within clades of one event. These findings indicate multiple and separate introductions of cowpox virus, probably from wild rodent populations, where the virus keeps circulating naturally and is only sporadically introduced into the cheetahs. Sero-positivity of voles (Arvicola amphibious) caught in zoo grounds strengthens this hypothesis. As a consequence, recommendations are given for medical and physical management of diseased cheetahs, for hygienic measures as well as for pre-shipment isolation before cheetah export from zoo grounds.

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“…Since 2001, real-time PCRs have been developed (e.g. Czerny et al 2001;Kulesh et al 2004;Balamurugan et al 2009;Gavrilova et al 2010;Li et al 2010;Shchelkunov et al 2011;Venkatesan et al 2012a). A DNA oligonucleotide microarray with the TNF receptor gene crmB (Lapa et al 2002) and loopmediated isothermal amplification (LAMP) assays are alternative rapid methods for OPXV detection or differentiation (Iizuka et al 2009;Venkatesan et al 2012b).…”

Section: Diagnosismentioning

confidence: 99%