Western blotting analysis of <i>L</i>. <i>lactis</i> MG1363 PPiA KO mutant and its complemented mutant, where the SpaA-SrtC1 cassette was introduced.

<p>SpaA pilin proteins were detected using anti-SpaA polyclonal antibodies. Legend: #, protein weight marker; HMWL, high molecular weight ladder; *, band corresponding to elongated pilus structures.</p

Bacterial strains and plasmids used in the present study.

<p>Bacterial strains and plasmids used in the present study.</p

Summary of pilus phenotypes observed in derivatives of <i>Lactococcus lactis</i> NZ9000, <i>Lactobacillus rhamnosus</i> GG and PB12.

<p>The AA replacement is shaded in light grey within the GYPSY motif (bold AA residues).</p

Western blotting analysis of <i>L</i>. <i>lactis</i> NZ9000 expressing different <i>spaA-srtC</i>1 gene variants, where the GYPSY motif has been altered by AA substitution.

<p><b>SpaA pilin proteins were detected using anti-SpaA polyclonal antibodies.</b> Legend: #, protein weight marker; HMWL, high molecular weight ladder; *, band corresponding to elongated pilus structures.</p

Electron microscopy observations of <i>Lb</i>. <i>rhamnosus</i> PB12 and its SrtC1-complemented derivatives.

<p>Bacterial cells were immunogold-labeled with anti-SpaA serum and gold particles (10 nm). Legend: A, <i>Lb</i>. <i>rhamnosus</i> GG; B, <i>Lb</i>. <i>rhamnosus</i> PB12; C, <i>Lb</i>. <i>rhamnosus</i> PB12 + SrtC1.</p

Electron microscopy observations of <i>L</i>. <i>lactis</i> NZ9000 producing various SpaA pilus structures.

<p>Bacterial cells were immunogold-labeled with anti-SpaA serum and gold particles (10 nm). Legend: A, NZ9000 + SpaA-SrtC1; B, NZ9000 + SpaA-SrtC1^{Y29G, P30G}; C, NZ9000 + SpaA-SrtC1^P30G; D, NZ9000 + SpaA-SrtC1^Y29G.</p

Modelling of the region containing the GYPSY motif of the <i>Lb</i>. <i>rhamnosus</i> GG SrtC1 with kinked proline residue.

<p>(A) Output prediction results using PSIPRED server [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0153373#pone.0153373.ref043" target="_blank">43</a>,<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0153373#pone.0153373.ref044" target="_blank">44</a>] indicated that the GYPSY motif introduces a breakage within an α-helix as opposed to some mutant variants; (B) and (C), <i>ab initio</i> model of the GYPSY motif. The kinked proline residue is shaded in fuchsia. The relevant AA modification(s) introduced in the present study are shown in vivid green.</p

Defense systems of the multi-species probiotic product VSL#3.

<p>Number of predicted restriction/modification enzymes (blue) and CRISPR-Cas spacers (green). Strains devoid of CRISPR-Cas loci have a number of spacers equal to zero.</p

CRISPR-Cas loci identified in the strains of the VSL#3 product.

<p>The CRISPR-Cas loci were identified using CRISPRFinder [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0192452#pone.0192452.ref056" target="_blank">56</a>] and their gene order and predicted annotations are depicted along with their juxtaposing array of spacers. Legend: *, split gene.</p

Mucus binding assays of <i>Lb</i>. <i>rhamnosus</i> GG and PB12 expressing different <i>srtC</i>1 gene variants, where the GYPSY motif has been altered by AA substitution.

<p>The averages and standard deviations were obtained from a total of twelve technical replicates from three biological replicates.</p