Fig. 3: Activity of defense systems against phages infecting S. thermophilus and L. cremoris.

A Strategy for assessing the efficiency of defense systems against streptococcal phages from five genera and lactococcal phages from two genera. Anti-phage activity was measured by evaluating the EOP of phages on the host strain containing the defense mechanism cloned on a plasmid compared to the host strain with the empty plasmid. B Comparison of Dodola and Kiwa defense systems efficiency against L. cremoris phage p2 when cloned into low-copy and high-copy vectors. C EOP heatmap of phages on strains carrying each defense system. Darker shades indicate stronger anti-phage protection. An EOP of 10⁻¹ was used as the minimum threshold for anti-phage activity. Three biological replicates were done. Some defense systems were not tested against a few phages, as they are naturally present in corresponding host strains. Two homologs were tested for Gao19, PD-Lambda-1, RosmerTA, and Hachiman. Phages encoding known ACR are highlighted in bold. The top to bottom presentation of the defense systems reflects the high to low prevalence (Fig. 1A). The genetic organization of the defense systems is shown on the right side of the heatmap. A detailed description of their functional domains is available in Supplementary Data 6. D Comparison of anti-phage activity of two Gao19 homologs. Gao19 was found in 35 S. thermophilus strains, of which 2 corresponded to Gao19^ST3 and 9 to Gao19^ST109. Bars show the mean EOP with error bars indicating standard deviation from biological replicates (n = 3). Filled circles indicate the presence of countable plaques, while hollow red circles denote lysis zones where plaques were not counted. Half circles indicate the absence of visible plaques or lysis zones even with undiluted phage lysate, showing that the defense system reduced phage levels below the limit of detection (EOP < 10⁻⁷). AB^R Antibiotic resistance gene, Cm^R Chloramphenicol resistance, DUF Domain of unknown function, EOP Efficiency of plaquing, Ery^R Erythromycin resistance, EV Empty vector, ori Origin of replication, SIR Sirtuin, TM Transmembrane domain. Source data are provided as a Source Data file.