Deletion of the rfaE gene in Haemophilus parasuis SC096 strain attenuates serum resistance, adhesion and invasion

• We generated a ΔrfaE mutant in Haemophilus parasuis SC096 strain.
• The ΔrfaE mutant exhibited truncated LOS structures and a pronounced growth defect.
• Loss of the rfaE gene resulted in more sensitivity in porcine and rabbit serum.
• The ΔrfaE mutant showed the impaired ability of adhering and invading on cells.

In Haemophilus parasuis, the lipooligosaccharide (LOS) has been identified as an important virulence factor. The rfa gene cluster encodes enzymes for LOS core biosynthesis. In order to investigate the role of the rfaE gene, we generated an rfaE deficient mutant (ΔrfaE) of a H. parasuis SC096 by a natural transformation method. The purified preparation of LOS from the ΔrfaE mutant strain showed truncated LOS structure on silver-stained SDS-PAGE. Compared to the wild-type SC096 strain, the generation time of ΔrfaE mutant strain was significantly extended from 59 min to 69 min. The ΔrfaE mutant strain caused an approximately 30-fold reductions in survival rate in 50% sera and 36-fold reductions in survival rate in 90% sera, respectively (p < 0.001). In adhesion and invasion assays, the ΔrfaE mutant strain had 10-fold less efficient adherence and 12-fold reductions in invasion of the porcine umbilicus vein endothelial cells (PUVEC) and porcine kidney epithelial cells (PK-15), respectively (p < 0.001). However, the complemented strain could restore the above phenotypes. Hence, the above results suggested that the rfaE gene participated in the pathogenicity of H. parasuis SC096 strain.