Role of the Genes of Type VI Secretion System in Virulence of Rice Bacterial Brown Stripe Pathogen Acidovorax avenae subsp. avenae Strain RS-2

Abstract

The Type VI secretion system (T6SS) is a class of macromolecular machine that is required for the virulence of gram-negative bacteria. However, it is still not clear what the role of T6SS in the virulence of rice bacterial brown stripe pathogen Acidovorax avenae subsp. avenae (Aaa) is. The aim of the current study was to investigate the contribution of T6SS in Aaa strain RS2 virulence using insertional deletion mutation and complementation approaches. This strain produced weak virulence but contains a complete T6SS gene cluster based on a genome-wide analysis. Here we compared the virulence-related phenotypes between the wild-type (RS-2) and 25 T6SS mutants, which were constructed using homologous recombination methods. The mutation of 15 T6SS genes significantly reduced bacterial virulence and the secretion of Hcp protein. Additionally, the complemented 7 mutations ΔpppA, ΔclpB, Δhcp, ΔdotU, ΔicmF, ΔimpJ, and ΔimpM caused similar virulence characteristics as RS-2. Moreover, the mutant ΔpppA, ΔclpB, ΔicmF, ΔimpJ and ΔimpM genes caused by a 38.3~56.4% reduction in biofilm formation while the mutants ΔpppA, ΔclpB, ΔicmF and Δhcp resulted in a 37.5~44.6% reduction in motility. All together, these results demonstrate that T6SS play vital roles in the virulence of strain RS-2, which may be partially attributed to the reductions in Hcp secretion, biofilm formation and motility. However, differences in virulence between strain RS-1 and RS-2 suggest that other factors may also be involved in the virulence of Aaa.

Halotolerant bacteria belonging to operational group Bacillus amyloliquefaciens in biocontrol of the rice brown stripe pathogen Acidovorax oryzae

“…Obviously, this rice pathogen have a harmful effect on seedlings height, which has been used for evaluating the virulence of Ao in our previous studies (Masum et al . ; Zhang et al . ).…”

“…The inhibitory effect of culture filtrates on bacterial growth may be, at least partially, attributed to the inhibition of virulence determinants such as swimming motility and biofilm formation, which has been found to be very closely associated with the virulence of Ao strains RS‐1 and RS‐2 in our previous studies (Masum et al . ; Zhang et al . ).…”

“…; Masum et al . ). Bacteria were cultured in Luria–Bertani (LB) (tryptone 10 g l −1 , yeast extract 5 g l −1 , NaCl 10 g l −1 , agar 15 g l −1 , pH 7·0) agar or broth medium (Sambrook et al .…”

“…The effect of culture filtrates on motility of Ao strains RS‐1 and RS‐2 was determined as described by Masum et al . (), which was performed by spotting pathogenic bacteria on the centre of each swimming LB plate (0·3% agar) supplemented with culture filtrate of antagonistic bacteria and then measuring the colony diameter after 48 h of incubation at 30°C. This experiment was repeated twice independently with three replications.…”

“…Two pathogenic strains RS-1 and RS-2 of Ao were used in this study, which were isolated from diseased rice plants in our previous studies Masum et al 2017). Bacteria were cultured in Luria-Bertani (LB) (tryptone 10 g l À1 , yeast extract 5 g l À1 , NaCl 10 g l À1 , agar 15 g l À1 , pH 7Á0) agar or broth medium (Sambrook et al 1989) at 30°C.…”