Development and application of an indirect ELISA and nested PCR for the epidemiological analysis of among pigs in China.

INTRODUCTION

() is an important opportunistic and zoonotic pathogen which is associated with many diseases in humans and animals. However, the pathogenicity of has been neglected and the prevalence of is poorly studied due to the lack of rapid and sensitive diagnosis techniques.

METHODS

In this study, we infected mice and pigs with strain from a human patient. An indirect ELISA was established using the KHE protein as the coating protein for the detection of specific antibody in clinical samples. A nested PCR method to detect nuclei acids of was also developed.

RESULTS

We showed that infection with strain from a human patient led to mild lung injury of pigs. For the ELISA, the optimal coating concentration of KHE protein was 10 µg/mL. The optimal dilutions of serum samples and secondary antibody were 1:100 and 1:2500, respectively. The analytical sensitivity was 1:800, with no cross-reaction between the coated antigen and porcine serum positive for antibodies against other bacteria. The intra-assay and inter-assay reproducibility coefficients of variation are less than 10%. Detection of 920 clinical porcine serum samples revealed a high infection rate by established indirect ELISA (27.28%) and nested PCR (19.13%). Moreover, correlation analysis demonstrated infection rate is positively correlated with gross population, Gross Domestic Product (GDP), and domestic tourists.

DISCUSSION

In conclusion, is highly prevalent among pigs in China. Our study highlights the role of in pig health, which provides a reference for the prevention and control of diseases associated with .