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Development of an Indirect Dot-PPA-ELISA using glutamate dehydrogenase as a diagnostic antigen for the rapid and specific detection of Streptococcus suis and its application to clinical specimens

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Abstract

Streptococcus suis is an important zoonotic pathogen causing infections in pigs and humans. Bacterial surface-related proteins are often explored as potential vaccine candidates and diagnostic antigens. In the present study, glutamate dehydrogenase, a highly conserved immunogenic extracellular protein, was used to establish a dot horseradish peroxidase enzyme-linked staphylococcal protein A immunosorbent assay (Dot-PPA-ELISA) for diagnosis of S. suis infection. The antigen–antibody reaction was optimised through checkerboard titration involving serial dilutions, followed by selective blocking tests and evaluations of cross-reaction, repeatability, and stability. Comparative analysis by using a conventional plate ELISA kit showed that the specificity and sensitivity of the Dot-PPA-ELISA were 97.5 and 96.6%, respectively. Furthermore, dynamic changes in the levels of antibody in rabbits immunised with a propolis inactivated vaccine were monitored by Dot-PPA-ELISA. A total seroprevalence of 73.1% in 305 pig serum samples indicated the method’s applicability to detect S. suis infection. Cumulatively, the results suggested that Dot-PAA-ELISA is a convenient, rapid, sensitive, and specific diagnostic method suitable for studying large numbers of samples obtained from clinical and epidemiological studies, thereby helping reduce important economic losses.

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Authors and Affiliations

  1. College of Animal Science and Veterinary Medicine, Shandong Agricultural University, No. 61, Daizong Street, Taian, 271018, Shandong, People’s Republic of China

    Xiao-jing Xia, Zhi-qiang Shen & Shi-jin Jiang

  2. College of Animal Science and Veterinary Medicine, Henan Institute of Science and Technology, No. 90, Hualan Street, Xinxiang, 453003, Henan, People’s Republic of China

    Xiao-jing Xia, Lei Wang & Jianhe Hu

  3. Shandong Binzhou Animal Science &Veterinary Medicine Academy, No.169, Huanghe 2th road, Binzhou, 256600, Shandong, People’s Republic of China

    Zhi-qiang Shen & Shu-guang Li

  4. Center for Experimental and Molecular Medicine, Academic Medical Center, University of Amsterdam, 1105 AZ, Amsterdam, The Netherlands

    Wanhai Qin

Authors
  1. Xiao-jing Xia
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  2. Lei Wang
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  3. Zhi-qiang Shen
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  4. Wanhai Qin
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  5. Jianhe Hu
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  6. Shi-jin Jiang
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  7. Shu-guang Li
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Corresponding authors

Correspondence to Zhi-qiang Shen or Shi-jin Jiang.

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10482_2016_825_MOESM1_ESM.tif

Supplementary material 1 (TIFF 1288 kb) Supplementary Fig. 1. Optimised concentration of antibody and antigen of the Indirect Dot-PPA-ELISA. Rows for sera of twofold dilutions from 1:10 to 1:2560, Row 10 for the control; lanes for the antigen of twofold dilutions from 0.3 µg/ml to 20 µg/ml, lane 8 for the control

10482_2016_825_MOESM2_ESM.tif

Supplementary material 2 (TIFF 1305 kb) Supplementary Fig. 2. Evaluation of assay repeatability of the Indirect Dot-PPA-ELISA. (a) Five positive serum samples were selected for intra-assay repeatability, three replicates of the same serum sample were performed in the same sheet. (b) Five positive serum samples were selected for inter-assay repeatability, three replicates of each sample were run in different sheets on different occasions.(c) Five batches of the rGDH protein were purified and used for coating antigen, three replicates of each batched were run in same sheets on same occasions

10482_2016_825_MOESM3_ESM.tif

Supplementary material 3 (TIFF 539 kb) Evaluation of assay stability of the Indirect Dot-PPA-ELISA. The coated NC sheet were stored at −20, 4, 25 or 37 °C for 1-6 month, and subsequently were used for the Indirect Dot-PPA-ELISA detection

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Xia, Xj., Wang, L., Shen, Zq. et al. Development of an Indirect Dot-PPA-ELISA using glutamate dehydrogenase as a diagnostic antigen for the rapid and specific detection of Streptococcus suis and its application to clinical specimens. Antonie van Leeuwenhoek 110, 585–592 (2017). https://doi.org/10.1007/s10482-016-0825-z

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  • DOI: https://doi.org/10.1007/s10482-016-0825-z

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