Csp1, a cold-shock protein homolog in Xylella fastidiosa is involved in stress response and biofilm formation

Bacterial cold shock-domain proteins (CSPs) are conserved nucleic acid binding chaperone proteins that play important roles in adaptation to environmental changes and stressors such as cold adaptation and conditions unrelated to temperature. Csp1, a temperature-independent cold shock protein homolog, acts as a virulence factor in Xylella fastidiosa, a bacterial pathogen of grapevine and other economically important crops. In addition to virulence, Csp1 contributes to X. fastidiosa survival at low temperatures and under high salt and oxidative stress conditions, in vitro. However, little is known about the specific function(s) of Csp1 in X. fastidiosa, aside from general single-stranded nucleic acid binding activity. To investigate the influence of Csp1 on X. fastidiosa gene expression, we used Nanopore’s MinION to sequence transcriptomes of wild-type X. fastidiosa strain Stag’s Leap and a csp1 deletion mutant (Δcsp1) under standard growth conditions (28°C) and cold stress (15°C). Our analysis revealed changes in expression of several genes important for motility and biofilm formation in Δcsp1 compared to wild-type. One gene of intertest, pilA1, encodes a type IV pili subunit protein and was up-regulated in the Δcsp1 mutant. The type IV pili is required for twitching motility in X. fastidiosa and contributes to virulence by facilitating bacterial movement inside the host vascular tissue. Previous studies in other strains of X. fastidiosa showed deleting pilA1 leads to overabundance of type IV pili and increased biofilm formation. Xylem vessel occlusion caused by biofilms is one major mechanism of X. fastidiosa pathogenicity, so Csp1 may act as a virulence factor by influencing the expression of genes involved in biofilm formation and motility. We also observed a significant decrease in biofilm formation with the Δcsp1 mutant compared to the wild type strain in liquid media, which may be the result of increased expression of pilA1.  The csp1 mutant was also less viable during long term growth compared to the wild-type strain, in vitro, further indicating Csp1 may be involved in stress response. Studies in other bacteria have identified links between stress response and biofilm formation, suggesting that Csp1 may play a role in both virulence and stress response by influencing genes important for biofilm formation.