Development of a sensitive and specific xMAP assay for detection of antibodies against infectious laryngotracheitis and bronchitis viruses

Wang, Huanan; Feng, Cong; Guan, Jianchi; Xiao, Li; Zhu, Yujun; Lian, Yuexiao; Ren, Haitao; Chen, Meili; Guo, Pengju

doi:10.1186/s12985-018-1048-x

Cited by 12 publications

(12 citation statements)

References 28 publications

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“…Based on the promising results obtained with the BBMA using human sera and the strong performance of TgSAG1 bio [37,39], we strived for a transfer of this assay to animal species, including chickens, to establish an improved method for large scale, efficient serological monitoring. Although there is a number of BBMAs for veterinary purposes, these tests mainly focus on viral infections [41][42][43][44] and cannot be easily compared with our assay, which, to the best of our knowledge represents the first BBMA focusing on parasitic pathogens in chickens.…”

Section: Resultsmentioning

confidence: 99%

Fluorescent bead-based serological detection of Toxoplasma gondii infection in chickens

et al. 2020

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Background: Free-ranging chickens are often infected with Toxoplasma gondii and seroconvert upon infection. This indicates environmental contamination with T. gondii. Methods: Here, we established a bead-based multiplex assay (BBMA) using the Luminex technology for the detection of T. gondii infections in chickens. Recombinant biotinylated T. gondii surface antigen 1 (TgSAG1 bio) bound to streptavidin-conjugated magnetic Luminex beads served as antigen. Serum antibodies were detected by a fluorophore-coupled secondary antibody. Beads of differing color codes were conjugated with anti-chicken IgY or chicken serum albumin and served for each sample as an internal positive or negative control, respectively. The assay was validated with sera from experimentally and naturally infected chickens. The results were compared to those from reference methods, including other serological tests, PCRs and bioassay in mice. Results: In experimentally infected chickens, the vast majority (98.5%, n = 65/66) of birds tested seropositive in the BBMA. This included all chickens positive by magnetic-capture PCR (100%, n = 45/45). Most, but not all inoculated and TgSAG1 bio-BBMA-positive chickens were also positive in two previously established TgSAG1-ELISAs (TgSAG1-ELISA SL , n = 61/65; or TgSAG1-ELISA SH , n = 60/65), or positive in an immunofluorescence assay (IFAT, n = 64/65) and in a modified agglutination test (MAT, n = 61/65). All non-inoculated control animals (n = 28/28, 100%) tested negative. In naturally exposed chickens, the TgSAG1 bio-BBMA showed a high sensitivity (98.5%; 95% confidence interval, CI: 90.7-99.9%) and specificity (100%; 95% CI: 85.0-100%) relative to a reference standard established using ELISA, IFAT and MAT. Almost all naturally exposed chickens that were positive in bioassay or by PCR tested positive in the TgSAG1 bio-BBMA (93.5%; 95% CI: 77.1-98.9%), while all bioassay-or PCR-negative chickens remained negative (100%; 95% CI: 85.0-100%). Conclusions: The TgSAG1 bio-BBMA represents a suitable method for the detection of T. gondii infections in chickens with high sensitivity and specificity, which is comparable or even superior to other tests. Since assays based on this methodology allow for the simultaneous analysis of a single biological sample with respect to multiple analytes, the described assay may represent a component in future multiplex assays for broad serological monitoring of poultry and other farm animals for various pathogens.

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Section: Resultsmentioning

confidence: 99%

Fluorescent bead-based serological detection of Toxoplasma gondii infection in chickens

et al. 2020

View full text Add to dashboard Cite

show abstract

“…Based on the promising results obtained with the BBMA using human sera and the strong performance of TgSAG1 bio [37,39], we strived for a transfer of this assay to other animal species, including chickens, to establish an improved method for large scale, e cient serological monitoring. Although there is a number of BBMAs for veterinary purposes, these tests mainly focus on viral infections [41,42,43,44] and cannot be easily compared with our assay, which represents the rst BBMA focusing on parasitic pathogens in chickens.…”

Section: Resultsmentioning

confidence: 99%

Fluorescent bead-based serological detection of Toxoplasma gondii infection in chickens

Fabian

Hedar

Koethe

et al. 2020

Preprint

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Background: Free-ranging chickens are often infected with Toxoplasma gondii. Their infection indicates environmental contamination with T. gondii. The detection of infected birds relies primarily on serological assays. Methods: Here, we established a bead-based multiplex assay (BBMA) using the Luminex technology for the specific and sensitive detection of T. gondii infections in chickens. Recombinant biotinylated T. gondii surface antigen 1 (TgSAG1bio) bound to streptavidin-conjugated magnetic Luminex beads served as antigen. Serum antibodies were detected by a fluorophore-coupled secondary antibody. Beads of differing color codes were conjugated with anti-chicken IgY or chicken serum albumin and served for each sample as an internal positive or negative control, respectively. The assay was validated with sera from experimentally and naturally infected chickens. The results were compared to those from reference methods, including other serological tests and bioassay in mice.Results: In experimentally infected chickens, the vast majority (98.5%, n=65/66) of inoculated birds tested seropositive in the BBMA. This included all chickens positive by magnetic-capture PCR (100%, n=45/45). Most, but not all inoculated and TgSAG1bio-BBMA-positive chickens were also positive in two previously established TgSAG1-ELISAs (TgSAG1-ELISASL, n=61/65; or TgSAG1-ELISASH, n=60/65), or positive in an immunofluorescence assay (IFAT, n=64/65)) and in a modified agglutination test (MAT, n=61/65). All non-inoculated control animals (n=28/28, 100%) tested negative. In naturally exposed chickens, the TgSAG1bio-BBMA showed a high sensitivity (98.5%; 95% Confidence Interval: 90.7-99.9%) and specificity (100%; 85.0-100%) relative to a reference standard established using ELISA, IFAT and MAT. Almost all naturally exposed chickens that were positive in bioassay or by PCR tested positive in the TgSAG1bio-BBMA (93.5%; 77.1-98.9%), while all bioassay- or PCR-negative chickens remained negative (100%; 85.0-100%).Conclusions: The TgSAG1bio-BBMA represents a suitable method for the detection of T. gondii infections in chickens with high sensitivity and specificity, which is comparable or even superior to other tests. Since assays based on this methodology allow for the simultaneous analysis of a single biological sample with respect to multiple analytes, the described assay may represent one component in future multiplex assays for broad serological monitoring of poultry and other farm animals, including pigs or small ruminants, for various pathogens.

show abstract

“…Based on the promising results obtained with the BBMA using human sera and the strong performance of TgSAG1 bio [37,39], we strived for a transfer of this assay to other animal species, including chickens, to establish an improved method for large scale, e cient serological monitoring. Although there is a number of BBMAs for veterinary purposes, these tests mainly focus on viral infections [41][42][43][44] and cannot be easily compared with our assay, which, to the best of our knowledge represents the rst BBMA focusing on parasitic pathogens in chickens.…”

Section: Resultsmentioning

confidence: 99%

Fluorescent bead-based serological detection of Toxoplasma gondii infection in chickens

Fabian

Hedar

Koethe

et al. 2020

Preprint

View full text Add to dashboard Cite

Background: Free-ranging chickens are often infected with Toxoplasma gondii and seroconvert upon infection. This indicates environmental contamination with T. gondii. Methods: Here, we established a bead-based multiplex assay (BBMA) using the Luminex technology for the detection of T. gondii infections in chickens. Recombinant biotinylated T. gondii surface antigen 1 (TgSAG1bio) bound to streptavidin-conjugated magnetic Luminex beads served as antigen. Serum antibodies were detected by a fluorophore-coupled secondary antibody. Beads of differing color codes were conjugated with anti-chicken IgY or chicken serum albumin and served for each sample as an internal positive or negative control, respectively. The assay was validated with sera from experimentally and naturally infected chickens. The results were compared to those from reference methods, including other serological tests, PCRs and bioassay in mice.Results: In experimentally infected chickens, the vast majority (98.5%, n = 65/66) of birds tested seropositive in the BBMA. This included all chickens positive by magnetic-capture PCR (100%, n = 45/45). Most, but not all inoculated and TgSAG1bio-BBMA-positive chickens were also positive in two previously established TgSAG1-ELISAs (TgSAG1-ELISASL, n = 61/65; or TgSAG1-ELISASH, n = 60/65), or positive in an immunofluorescence assay (IFAT, n = 64/65)) and in a modified agglutination test (MAT, n = 61/65). All non-inoculated control animals (n = 28/28, 100%) tested negative. In naturally exposed chickens, the TgSAG1bio-BBMA showed a high sensitivity (98.5%; 95% confidence interval, CI: 90.7–99.9%) and specificity (100%; 95% CI: 85.0–100%) relative to a reference standard established using ELISA, IFAT and MAT. Almost all naturally exposed chickens that were positive in bioassay or by PCR tested positive in the TgSAG1bio-BBMA (93.5%; 95% CI: 77.1–98.9%), while all bioassay- or PCR-negative chickens remained negative (100%; 95% CI: 85.0–100%).Conclusions: The TgSAG1bio-BBMA represents a suitable method for the detection of T. gondii infections in chickens with high sensitivity and specificity, which is comparable or even superior to other tests. Since assays based on this methodology allow for the simultaneous analysis of a single biological sample with respect to multiple analytes, the described assay may represent a component in future multiplex assays for broad serological monitoring of poultry and other farm animals for various pathogens.

show abstract

Development of a sensitive and specific xMAP assay for detection of antibodies against infectious laryngotracheitis and bronchitis viruses

Cited by 12 publications

References 28 publications

Fluorescent bead-based serological detection of Toxoplasma gondii infection in chickens

Fluorescent bead-based serological detection of Toxoplasma gondii infection in chickens

Fluorescent bead-based serological detection of Toxoplasma gondii infection in chickens

Fluorescent bead-based serological detection of Toxoplasma gondii infection in chickens

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