Journal: PLOS Genetics

Article Title: Feature architecture aware phylogenetic profiling indicates a functional diversification of type IVa pili in the nosocomial pathogen Acinetobacter baumannii

doi: 10.1371/journal.pgen.1010646

Figure Lengend Snippet: (A) Representative electron micrograph of wild-type A . baumannii AYE-T cells (top) and of the Δ comC mutant (bottom). No piliation defect is seen for the mutant. A phenotyping of wild-type A . baumannii AYE-T, of the Δ comC mutant, and of the mutant complemented with the indicated constructs are shown in the following panels. (B) Adhesion to HUVEC cells. The bar height and error bar indicate the mean number and standard deviation, respectively, of colony forming units (CFU) per mL (n = 4; values are given as ). ‘*’ indicates a significant difference (one tailed t test: p<0.05). Complementation with the truncated comC mutants did not significantly increase adhesion rates. (C) Twitching motility. Cells were stained with 1% [w/v] crystal violet. (D) Natural transformation rates of the indicated A . baumannii cells were assessed using genomic DNA of rifampicin resistant A . baumannii ATCC 19606 T . n.d.—not detected.

Article Snippet: Three different constructs were used to complement Ab AYE-T ΔcomC :: kanR : (i) full length ComC–The comC gene plus 700 bp of the upstream region were amplified from chromosomal DNA of A . baumannii ATCC 19606 T using primers 11–12 ( ).

Techniques: Mutagenesis, Construct, Standard Deviation, One-tailed Test, Staining, Transformation Assay

Journal: PLOS Genetics

Article Title: The novel anti-phage system Shield co-opts an RmuC domain to mediate phage defense across Pseudomonas species

doi: 10.1371/journal.pgen.1010784

Figure Lengend Snippet: (a) Cells carrying plasmid pBAD18 encoding ShdA II-His 6 were grown for 5 hrs in the presence of 0.2% L-arabinose. Cells were fractionated to produce soluble and membrane samples and analysed by immunoblot with antibodies to the His6 tag, GroEL (cytoplasmic control) and TatA (membrane control). (b-e) In vitro DNAse activity assays using (b) ShdA II 138-524 (c) ShdA I 140-526 (d) ShdA III 135-523 and (e) ShdA IV 135-521 . ShdA proteins and DHFR were synthesised using the cell-free PURExpress kit (NEB). DNAse activity was tested against 10 ng of input DNA. DNA types tested were phage DNA, E . coli MG1655 chromosomal DNA and plasmid (pSG483) DNA. For ShdA I 140-526 , ShdA III 135-523 and ShdA II 138-524 phage DNA was from ϕSipho. For ShdA IV 135-521 phage DNA was from ϕAlma. (f) Evaluation of transformation efficiency for E . coli MG1655 carrying empty vector (VC, pBAD18) or the same plasmid encoding Shield subtype II with plasmid DNA. For all panels, 0.2% L-arabinose was added at time zero to induce expression of the encoded genes in pBAD18. Points show mean +/− SEM (n = 3 biological replicates). Statistical analysis was performed using one-way ANOVA with Dunnett’s multiple comparison test. No significance was detected, unless indicated (*p ≤ 0.05).

Article Snippet: The strain P . aeruginosa NCTC 11442 (equivalent to ATCC 33350) was grown overnight at 37°C for chromosomal DNA extraction, whereas of P . aeruginosa PA14 csy3 :: lacZ was grown at 30°C.

Techniques: Plasmid Preparation, Western Blot, In Vitro, Activity Assay, Transformation Assay, Expressing