Screening for Virulence-Related Genes via a Transposon Mutant Library of Streptococcus suis Serotype 2 Using a Galleria mellonella Larvae Infection Model

Streptococcus suis (S. suis) is a zoonotic bacterial pathogen causing lethal infections in pigs and humans. Identification of virulence-related genes (VRGs) is of great importance in understanding the pathobiology of a bacterial pathogen. To identify novel VRGs, a transposon (Tn) mutant library of S. suis strain SC19 was constructed in this study. The insertion sites of approximately 1700 mutants were identified by Tn-seq, which involved 417 different genes. A total of 32 attenuated strains were identified from the library by using a Galleria mellonella larvae infection model, and 30 novel VRGs were discovered, including transcription regulators, transporters, hypothetical proteins, etc. An isogenic deletion mutant of hxtR gene (Delta hxtR) and its complementary strain (C Delta hxtR) were constructed, and their virulence was compared with the wild-type strain in G. mellonella larvae and mice, which showed that disruption of hxtR significantly attenuated the virulence. Moreover, the Delta hxtR strain displayed a reduced survival ability in whole blood, increased sensitivity to phagocytosis, increased chain length, and growth defect. Taken together, this study performed a high throughput screening for VRGs of S. suis using a G. mellonella larvae model and further characterized a novel critical virulence factor.