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In vitro detection of prionemia in TSE-infected cervids and hamsters.

Elder AM, Henderson DM, Nalls AV, Wilham JM, Caughey BW, Hoover EA, Kincaid AE, Bartz JC, Mathiason CK - PLoS ONE (2013)

Bottom Line: Thus, it is important to monitor cervid and human blood products to ensure herd health and human safety.Current methods for detecting blood-associated prions rely primarily upon bioassay in laboratory animals.Our results indicate that RT-QuIC methodology as modified can provide consistent and reliable detection of blood-borne prions in preclinical and symptomatic stages of two animal TSEs, offering promise for prionemia detection in other species, including humans.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, Immunology and Pathology, Colorado State University, Fort Collins, Colorado, United States of America.

ABSTRACT
Blood-borne transmission of infectious prions during the symptomatic and asymptomatic stages of disease occurs for both human and animal transmissible spongiform encephalopathies (TSEs). The geographical distribution of the cervid TSE, chronic wasting disease (CWD), continues to spread across North America and the prospective number of individuals harboring an asymptomatic infection of human variant Creutzfeldt-Jakob Disease (vCJD) in the United Kingdom has been projected to be ~1 in 3000 residents. Thus, it is important to monitor cervid and human blood products to ensure herd health and human safety. Current methods for detecting blood-associated prions rely primarily upon bioassay in laboratory animals. While bioassay provides high sensitivity and specificity, it requires many months, animals, and it is costly. Here we report modification of the real time quaking-induced conversion (RT-QuIC) assay to detect blood-borne prions in whole blood from prion-infected preclinical white-tailed deer, muntjac deer, and Syrian hamsters, attaining sensitivity of >90% while maintaining 100% specificity. Our results indicate that RT-QuIC methodology as modified can provide consistent and reliable detection of blood-borne prions in preclinical and symptomatic stages of two animal TSEs, offering promise for prionemia detection in other species, including humans.

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RT-QuIC analysis of hamster whole blood samples.Blood samples were diluted to 10-2 and 8 replicates were analyzed over 2 experiments of 60 hours, and positivity was determined by ThT fluorescence level above threshold. PrPC-converting activity is demonstrated in 21 TME-infected blood samples, and is absent in all TME-naïve samples (A-D). Each line is the average of four replicates for a specific animal. UN= Uninfected; INF= Infected.
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pone-0080203-g006: RT-QuIC analysis of hamster whole blood samples.Blood samples were diluted to 10-2 and 8 replicates were analyzed over 2 experiments of 60 hours, and positivity was determined by ThT fluorescence level above threshold. PrPC-converting activity is demonstrated in 21 TME-infected blood samples, and is absent in all TME-naïve samples (A-D). Each line is the average of four replicates for a specific animal. UN= Uninfected; INF= Infected.

Mentions: The hyper strain of transmissible mink encephalopathy (HY TME) was chosen for the RT-QuIC assay to determine the assays ability for PrPD detection in various species and strains of TSEs. All HY TME-infected hamsters (n=21), ranging from 8 to 20 weeks post infection, exhibited RT-QuIC PrPC-converting activity in 5/8 - 8/8 replicates within 60 hours, while all (n=7) of the age matched controls failed to seed RT-QuIC (Figure 6, Table 2). As above, each sample was run two separate times to determine consistency of the RT-QuIC assay. Sample replicates were averaged on each plate and a positive threshold was set at five times the standard deviation of the negative control average.


In vitro detection of prionemia in TSE-infected cervids and hamsters.

Elder AM, Henderson DM, Nalls AV, Wilham JM, Caughey BW, Hoover EA, Kincaid AE, Bartz JC, Mathiason CK - PLoS ONE (2013)

RT-QuIC analysis of hamster whole blood samples.Blood samples were diluted to 10-2 and 8 replicates were analyzed over 2 experiments of 60 hours, and positivity was determined by ThT fluorescence level above threshold. PrPC-converting activity is demonstrated in 21 TME-infected blood samples, and is absent in all TME-naïve samples (A-D). Each line is the average of four replicates for a specific animal. UN= Uninfected; INF= Infected.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3815098&req=5

pone-0080203-g006: RT-QuIC analysis of hamster whole blood samples.Blood samples were diluted to 10-2 and 8 replicates were analyzed over 2 experiments of 60 hours, and positivity was determined by ThT fluorescence level above threshold. PrPC-converting activity is demonstrated in 21 TME-infected blood samples, and is absent in all TME-naïve samples (A-D). Each line is the average of four replicates for a specific animal. UN= Uninfected; INF= Infected.
Mentions: The hyper strain of transmissible mink encephalopathy (HY TME) was chosen for the RT-QuIC assay to determine the assays ability for PrPD detection in various species and strains of TSEs. All HY TME-infected hamsters (n=21), ranging from 8 to 20 weeks post infection, exhibited RT-QuIC PrPC-converting activity in 5/8 - 8/8 replicates within 60 hours, while all (n=7) of the age matched controls failed to seed RT-QuIC (Figure 6, Table 2). As above, each sample was run two separate times to determine consistency of the RT-QuIC assay. Sample replicates were averaged on each plate and a positive threshold was set at five times the standard deviation of the negative control average.

Bottom Line: Thus, it is important to monitor cervid and human blood products to ensure herd health and human safety.Current methods for detecting blood-associated prions rely primarily upon bioassay in laboratory animals.Our results indicate that RT-QuIC methodology as modified can provide consistent and reliable detection of blood-borne prions in preclinical and symptomatic stages of two animal TSEs, offering promise for prionemia detection in other species, including humans.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, Immunology and Pathology, Colorado State University, Fort Collins, Colorado, United States of America.

ABSTRACT
Blood-borne transmission of infectious prions during the symptomatic and asymptomatic stages of disease occurs for both human and animal transmissible spongiform encephalopathies (TSEs). The geographical distribution of the cervid TSE, chronic wasting disease (CWD), continues to spread across North America and the prospective number of individuals harboring an asymptomatic infection of human variant Creutzfeldt-Jakob Disease (vCJD) in the United Kingdom has been projected to be ~1 in 3000 residents. Thus, it is important to monitor cervid and human blood products to ensure herd health and human safety. Current methods for detecting blood-associated prions rely primarily upon bioassay in laboratory animals. While bioassay provides high sensitivity and specificity, it requires many months, animals, and it is costly. Here we report modification of the real time quaking-induced conversion (RT-QuIC) assay to detect blood-borne prions in whole blood from prion-infected preclinical white-tailed deer, muntjac deer, and Syrian hamsters, attaining sensitivity of >90% while maintaining 100% specificity. Our results indicate that RT-QuIC methodology as modified can provide consistent and reliable detection of blood-borne prions in preclinical and symptomatic stages of two animal TSEs, offering promise for prionemia detection in other species, including humans.

Show MeSH
Related in: MedlinePlus