Csp1, a Cold Shock Protein Homolog in Xylella fastidiosa Influences Cell Attachment, Pili Formation, and Gene Expression.

Bacterial cold shock-domain proteins are conserved nucleic acid binding chaperones that play important roles in stress adaptation and pathogenesis. Csp1 is a temperature-independent cold shock protein homolog in Xylella fastidiosa, a bacterial plant pathogen of grapevine and other economically important crops. Csp1 contributes to stress tolerance and virulence in X. fastidiosa. However, besides general single-stranded nucleic acid binding activity, little is known about the specific function(s) of Csp1. To further investigate the role(s) of Csp1, we compared phenotypic differences and transcriptome profiles between the wild type and a deletion mutant (Δ). Csp1 contributes to attachment and long-term survival and influences gene expression. We observed reduced cell-to-cell attachment and reduced attachment to surfaces with the Δ strain compared to those with the wild type. Transmission electron microscopy imaging revealed that Δ was deficient in pili formation compared to the wild type and complemented strains. The Δ strain also showed reduced survival after long-term growth . Long-read nanopore transcriptome sequencing (RNA-Seq) analysis revealed changes in expression of several genes important for attachment and biofilm formation in Δ compared to that in the wild type. One gene of interest, , which encodes a type IV pili subunit protein, was upregulated in Δ. Deleting in X. fastidiosa strain Stag's Leap increased surface attachment and reduced virulence in grapevines. X. fastidiosa virulence depends on bacterial attachment to host tissue and movement within and between xylem vessels. Our results show that the impact of Csp1 on virulence may be due to changes in expression of attachment genes. Xylella fastidiosa is a major threat to the worldwide agriculture industry. Despite its global importance, many aspects of X. fastidiosa biology and pathogenicity are poorly understood. There are currently few effective solutions to suppress X. fastidiosa disease development or eliminate bacteria from infected plants. Recently, disease epidemics due to X. fastidiosa have greatly expanded, increasing the need for better disease prevention and control strategies. Our studies show a novel connection between cold shock protein Csp1 and pili abundance and attachment, which have not been reported for X. fastidiosa. Understanding how pathogenesis-related gene expression is regulated can aid in developing novel pathogen and disease control strategies. We also streamlined a bioinformatics protocol to process and analyze long-read nanopore bacterial RNA-Seq data, which will benefit the research community, particularly those working with non-model bacterial species.