Modified multiplex PCR for serotyping and pathotyping of Streptococcus suis

Introduction. Streptococcus suis is a zoonotic pathogen that causes invasive infections in humans who have been in close contact with infected pigs or contaminated pork-derived products. There is currently no consensus on the universal virulence factors or markers that can differentiate pathogenic from non-pathogenic or commensal S. suis isolates.Gap statement. A diagnostic tool for serotyping and pathotyping of S. suis is required for active public health surveillance and the One-Health approach.Aim. To improve the former multiplex PCR to serotyping all 29 recognized 'true' serotypes and distinguish pathogenic pathotypes using primers targeting the capsule and ROK pathogenic marker genes.Methodology. Four sets of multiplex PCRs were modified and improved to detect all 29 recognized serotypes of S. suis and distinguish their pathogenic pathotypes using the ROK gene.Results. This multiplex PCR allowed for the simultaneous amplification of S. suis-specific, serotype-specific and pathogenic pathotypes from the DNA of each serotype in each reaction. The accuracy, sensitivity, specificity, positive predictive value and negative predictive value of the pathogenic ROK marker genes were 84.7% (625/738), 96.4% (423/439), 67.6% (202/299), 81.4% (423/520) and 92.7% (202/218), respectively. There was a significant (P-value <0.001), high positive likelihood ratio [2.9 with 2.5-3.5 of 95% confidence interval (CI)] and a significant odds ratio (55.1 with 31.6-95.9 of 95 % CI), which indicated that the ROK gene could be used as the pathogenic pathotype marker. No cross-reactions were observed with other bacterial species.Conclusion. This modified multiplex PCR was able to distinguish 29 well-known serotypes and predicted the pathogenic pathotypes of S. suis isolates from humans and pigs in a single assay. It is useful for One-Health surveillance of human and pig isolates of S. suis.