Evaluating the most appropriate pooling ratio for EDTA blood samples to detect Bluetongue virus using real-time RT-PCR

Flannery, John; Rajko-Nenow, Paulina; Hicks, Hayley; Hill, Holly; Gubbins, Simon; Batten, Carrie

doi:10.1016/j.vetmic.2018.03.001

Cited by 9 publications

(11 citation statements)

References 25 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The vaccine-derived BTV-14 represents the most recent example of improper vaccination having an impact within Europe [16]. The infection kinetics of the BTV-14 strain (POL2012/01) used in our study were similar to those of other BTV serotypes in sheep [22][23][24]. In accordance with epidemiological findings in the field, BTV-14 elicited mostly mild clinical symptoms in the sheep species we used.…”

Section: Discussionsupporting

confidence: 82%

BTV-14 Infection in Sheep Elicits Viraemia with Mild Clinical Symptoms

et al. 2020

Self Cite

View full text Add to dashboard Cite

In 2011, Bluetongue virus serotype 14 (BTV-14) was detected in Russia during routine surveillance, and was subsequently found in a number of European countries. The strain had high sequence similarity to a BTV-14 vaccine strain. We aimed to determine the risk of this BTV-14 strain causing disease in a UK sheep breed. Four Poll Dorset sheep were infected with a Polish isolate of BTV-14 and infection kinetics were monitored over 28 days. BTV RNA was detected in EDTA blood by 4 days post-infection (dpi) and remained detectable at 28 days post-infection (dpi). Peak viraemia occurred at 6 and 7 dpi with Ct values ranging between 24.6 and 27.3 in all infected animals. BTV antibodies were detected by 10 dpi using a commercial ELISA and neutralising antibodies were detected from 10 dpi. BTV was isolated between 6 and 12 dpi. All infected sheep developed mild clinical signs such as reddening of conjunctiva and mucosal membranes, with one sheep demonstrating more overt clinical signs. Two uninoculated control animals remained clinically healthy and did not have detectable BTV RNA or antibodies. The overall mild clinical symptoms caused by this BTV-14 in this highly susceptible sheep breed were in accordance with the asymptomatic infections observed in the affected countries.

show abstract

Section: Discussionsupporting

confidence: 82%

BTV-14 Infection in Sheep Elicits Viraemia with Mild Clinical Symptoms

et al. 2020

Self Cite

View full text Add to dashboard Cite

show abstract

“…BTV RNA was extracted from 100 μl of EDTA blood and eluted into 80 μl buffer using the KingFisher Flex automated extraction platform and the MagVet Universal nucleic acid extraction kit (ThermoFisher Scientific, Paisley, UK). Ten microliters of sample RNA was analyzed as per the assay described by (Hofmann et al., ) with modifications (Flannery et al., ) using the Express One‐Step qRT‐PCR kit (ThermoFisher) on an Applied Biosystems 7500 Fast instrument (ThermoFisher). A log‐dilution series of the plasmid (1 × 10 0 –1 × 10 6 copies per μl) was included in triplicate on each RT‐qPCR run.…”

Section: Methodsmentioning

confidence: 99%

Evidence of reduced viremia, pathogenicity and vector competence in a re‐emerging European strain of bluetongue virus serotype 8 in sheep

Flannery

Sanz-Bernardo

Ashby

et al. 2019

Transbound Emerg Dis

Self Cite

View full text Add to dashboard Cite

Summary The outbreak of bluetongue virus (BTV) serotype 8 (BTV‐8) during 2006–2009 in Europe was the most costly epidemic of the virus in recorded history. In 2015, a BTV‐8 strain re‐emerged in France which has continued to circulate since then. To examine anecdotal reports of reduced pathogenicity and transmission efficiency, we investigated the infection kinetics of a 2007 UK BTV‐8 strain alongside the re‐emerging BTV‐8 strain isolated from France in 2017. Two groups of eight BTV‐naïve British mule sheep were inoculated with 5.75 log 10 TCID 50 /ml of either BTV‐8 strain. BTV RNA was detected by 2 dpi in both groups with peak viraemia occurring between 5–9 dpi. A significantly greater amount of BTV RNA was detected in sheep infected with the 2007 strain (6.0–8.8 log 10 genome copies/ml) than the re‐emerging BTV‐8 strain (2.9–7.9 log 10 genome copies/ml). All infected sheep developed BTV‐specific antibodies by 9 dpi. BTV was isolated from 2 dpi to 12 dpi for 2007 BTV‐8‐inoculated sheep and from 5 to 10 dpi for sheep inoculated with the remerging BTV‐8. In Culicoides sonorensis feeding on the sheep over the period 7–12 dpi, vector competence was significantly higher for the 2007 strain than the re‐emerging strain. Both the proportion of animals showing moderate (as opposed to mild or no) clinical disease (6/8 vs. 1/8) and the overall clinical scores (median 5.25 vs. 3) were significantly higher in sheep infected with the 2007 strain, compared to those infected with the re‐emerging strain. However, one sheep infected with the re‐emerging strain was euthanized at 16 dpi having developed severe lameness. This highlights the potential of the re‐emerging BTV‐8 to still cause illness in naïve ruminants with concurrent costs to the livestock industry.

show abstract

“…BTV RNA was extracted from 100 μL of EDTA blood and eluted into 80 μl buffer using the KingFisher Flex automated extraction platform and the MagVet Universal nucleic acid extraction kit (ThermoFisher Scientific, Paisley, UK). Ten microliters of sample RNA was analyzed as per the assay described by (Hofmann et al, 2008) with modifications (Flannery et al, 2018) using the Express One-Step qRT-PCR kit (ThermoFisher) on an Applied Biosystems 7500 Fast instrument (ThermoFisher). A log-dilution series of the plasmid (1 × 10 0 to 1 × 10 6 copies per μL) was included in triplicate on each RT-qPCR run.…”

Section: Molecular Analysesmentioning

confidence: 99%

Evidence of reduced viremia, pathogenicity and vector competence in a re-emerging European strain of bluetongue virus serotype 8 in sheep

Flannery

Sanz-Bernardo

Ashby

et al. 2018

Preprint

Self Cite

View full text Add to dashboard Cite

SummaryThe outbreak of bluetongue virus (BTV) serotype 8 (BTV-8) during 2006-2009 in Europe was the most costly epidemic of the virus in recorded history. In 2015, a BTV-8 strain re-emerged in France which has continued to circulate since then. To examine anecdotal reports of reduced pathogenicity and transmission efficiency, we investigated the infection kinetics of a 2007 UK BTV-8 strain alongside the re-emerging BTV-8 strain isolated from France in 2017. Two groups of eight BTV-naïve British mule sheep were inoculated with 5.75 log10TCID50 ml−1 of either BTV-8 strain. BTV RNA was detected by 2 dpi in both groups with peak viremia occurring between 5-9 dpi. A significantly greater amount of BTV RNA was detected in sheep infected with the 2007 strain (6.0-8.8 log10 genome copies mL−1) than the re-emerging BTV-8 strain (2.9-7.9 log10 genome copies mL−1). All infected sheep developed BTV-specific antibodies by 9 dpi. BTV was isolated from 2 dpi to 12 dpi for 2007 BTV-8-inoculated sheep and from 5 to 10 dpi for sheep inoculated with the remerging BTV-8. In Culicoides sonorensis feeding on the sheep over the period 7-12 dpi, vector competence was significantly higher for the 2007 strain than the re-emerging strain. Both the proportion of animals showing moderate (as opposed to mild or no) clinical disease (6/8 vs 1/8) and the overall clinical scores (median 5.25 vs 3) were significantly higher in sheep infected with the 2007 strain, compared to those infected with the re-emerging strain. However, one sheep infected with the re-emerging strain was euthanized at 16 dpi having developed severe lameness. This highlights the potential of the re-emerging BTV-8 to still cause illness in naïve ruminants with concurrent costs to the livestock industry.SummaryThe re-emerging Bluetongue virus serotype 8 still presents a threat to naïve ruminants in Europe despite reduced virulence

show abstract

Evaluating the most appropriate pooling ratio for EDTA blood samples to detect Bluetongue virus using real-time RT-PCR

Abstract: HighlightsBTV is detectable up to 30 days post infection in ruminants.Pooling samples at ratios >1:10 is suitable for surveillance in BTV-endemic areas.Peak BTV viraemia occurs between 7–12 d.p.i.Pooling at ratios >1:10 is suitable when animals are sampled 7–10 days post import.

Cited by 9 publications

References 25 publications

BTV-14 Infection in Sheep Elicits Viraemia with Mild Clinical Symptoms

BTV-14 Infection in Sheep Elicits Viraemia with Mild Clinical Symptoms

Evidence of reduced viremia, pathogenicity and vector competence in a re‐emerging European strain of bluetongue virus serotype 8 in sheep

Evidence of reduced viremia, pathogenicity and vector competence in a re-emerging European strain of bluetongue virus serotype 8 in sheep

Contact Info

Product

Resources

About

Evaluating the most appropriate pooling ratio for EDTA blood samples to detect Bluetongue virus using real-time RT-PCR

Abstract: HighlightsBTV is detectable up to 30 days post infection in ruminants.Pooling samples at ratios >1:10 is suitable for surveillance in BTV-endemic areas.Peak BTV viraemia occurs between 7–12 d.p.i.Pooling at ratios >1:10 is suitable when animals are sampled 7–10 days post import.

Cited by 9 publications

References 25 publications

BTV-14 Infection in Sheep Elicits Viraemia with Mild Clinical Symptoms

BTV-14 Infection in Sheep Elicits Viraemia with Mild Clinical Symptoms

Evidence of reduced viremia, pathogenicity and vector competence in a re‐emerging European strain of bluetongue virus serotype 8 in sheep

Evidence of reduced viremia, pathogenicity and vector competence in a re-emerging European strain of bluetongue virus serotype 8 in sheep

Contact Info

Product

Resources

About

Abstract: HighlightsBTV is detectable up to 30 days post infection in ruminants.Pooling samples at ratios >1:10 is suitable for surveillance in BTV-endemic areas.Peak BTV viraemia occurs between 7–12 d.p.i.Pooling at ratios >1:10 is suitable when animals are sampled 7–10 days post import.