Journal: Microorganisms

Article Title: Salmonella Heterogeneously Expresses Flagellin during Colonization of Plants

doi: 10.3390/microorganisms8060815

Figure Lengend Snippet: Salmonella persistence in and on leaves does not require flagella. Salmonella enterica serovar Typhimurium strain 14028s ( S . Typhimurium 14028s) and the double S . Typhimurium ΔfljBΔfliC mutant were infiltrated at OD 600 nm = 0.01 (ca. 10 7 CFU/mL) into tomato leaves. The presence of viable colony forming units (CFUs) was assessed during the following 14 days in cutout leaf discs. Both Salmonella strains persisted at the same level after infiltration ( a ). To monitor Salmonella growth in tomato-based (TM) medium, the number of CFU of S . Typhimurium 14028s and the double mutant S . Typhimurium ΔfljBΔfliC was assessed during 24 h post inoculation (hpi) into TM ( b ). Both strains reached the steady state during the 24 h, no difference between the wild type and the mutant proliferation rates was observed. Translocation of bacteria to non-inoculated leaves was verified using two inoculation techniques: infiltration ( c ) and dipping ( d ). After inoculation of leaves with bacterial suspensions (10 7 CFU/mL for infiltration or 10 8 CFU/mL for dipping), leaves were cut at the indicated days post inoculation (dpi) and subsequently incubated in enrichment media Buffered Peptone Water (BPW) and Rappaport Vassiliadis Broth (RVS). Graphs represent the translocation efficiency after infiltration ( c ) or dipping ( d ) inoculation. The enrichment cultures in RVS medium were plated on XLD-agar medium. The bars represent the percentage of plants for which non-inoculated leaves were positive for Salmonella from an average of seven (infiltration) and eight (dipping) replicates (one leaf per plant per treatment). ( e ) Direct comparison between persistence of S . Typhimurium 14028s and the double mutant S . Typhimurium ΔfljBΔfliC was assessed using competitive index (CI) assays. Both strains were co-inoculated by infiltrating a 5 × 10 5 CFU/mL with a 1:1 proportion of these two strains. Bacteria were extracted 7 and 14 dpi from tomato leaves and plated on LB plates for CFU determination. Replica plating was carried out in LB and LB plus kanamycin to differentiate between the two strains. CIs presented are representative results from six replicates. Mean CI values are shown. Errors bars represent standard error. Each CI was analyzed using a homoscedastic and 2-tailed Student’s t -test and the null hypothesis that mean index is not significantly different from 1, p < 0.05 were considered significant.

Article Snippet: Leaves (inoculated and non-inoculated) were sampled 5 h (0 dpi), 7 and 14 dpi, cut with sterile scissors and placed in 50 mL conical tubes containing 10 mL Buffered Peptone Water (BPW) (Carl Roth GmbH & Co. KG).

Techniques: Mutagenesis, Translocation Assay, Bacteria, Incubation, Comparison