Journal: Applied Microbiology and Biotechnology

Article Title: The novel antimicrobial peptide PXL150 in the local treatment of skin and soft tissue infections

doi: 10.1007/s00253-012-4439-8

Figure Lengend Snippet: Antimicrobial effect of PXL150 was evaluated in MMC 99 assay against a panel of Gram-positive and Gram-negative bacterial strains and yeast strains in 0.037 % BHI or 50 % h.i. SWF

Article Snippet: The microbicidal effect of PXL150 was tested against S. aureus (American Type Culture Collection (ATCC) 12600), MRSA (ATCC 33591), Streptococcus pyogenes (ATCC 12344), Propionibacterium acnes (ATCC 6919), Staphylococcus epidermidis (ATCC 12228), E. coli (ATCC 11775), P. aeruginosa (ATCC 15442), Klebsiella pneumoniae (Culture Collection, University of Gothenburg (CCUG) 59413, clinical isolate resistant to penicillins, cephalosporins, aztreonam and carbapenems, with the reference strain ATCC 13883), Acinetobacter baumannii (CCUG 58437, clinical isolate resistant to tobramycin, trimsulfa, ciprofloxacin, cefotaxim, ceftazidim, meropenem, pipera/tazobactam, with the reference strain ATCC 19606) and the yeast strains Candida albicans (ATCC 64549), Candida parapsilosis (ATCC 22019), Candida glabrata (CCUG 35267) and Candida krusei (ATCC 6258) using a minimal microbicidal concentration (MMC) assay.

Techniques:

Journal: Journal of Innate Immunity

Article Title: Histamine Produced by Gram-Negative Bacteria Impairs Neutrophil's Antimicrobial Response by Engaging the Histamine 2 Receptor

doi: 10.1159/000525536

Figure Lengend Snippet: Histamine produced by A. baumannii ATCC 17978 in G. mellonella larvae contributes to pathogenicity. G. mellonella (10 larvae per group) were inoculated, through the proleg, with 5.6 × 10 4 CFU or 1.1 × 10 5 CFU of A. baumannii ATCC 17978 or hdc::TOPO mutant. Larval survival was recorded after 24 h–54 h. a Photograph of larvae 18 h postinfection with A. baumannii ATCC 17978 or hdc::TOPO mutant with 5.6 × 10 4 CFU (left panel), or 1.1 × 10 5 CFU (right panel). b Kaplan-Meier survival analysis upon inoculation of G. mellonella larvae with A. baumannii ATCC 17978 or hdc::TOPO mutant at 5.6 × 10 4 CFU (left panel), or 1.1 × 10 5 CFU (right panel). c (left panel) G. mellonella larvae were inoculated with PBS (controls), 5.6 × 10 4 CFU of A. baumannii ATCC 17978, or hdc::TOPO mutant for 18 h after which six surviving larvae were collected. The haemolymph of the six larvae were pooled together. The concentration of histamine in the cell-free haemolymph was determined by ELISA. The data represent mean values ± SD of triplicate values. c (right panel) The haemolymph of ten larvae were pooled and hemocytes (1 × 10 7 ) were incubated with C. albicans (2 × 10 6 ) in the absence (dashed line) or presence (solid line) of histamine (10 −6 M). After 10–20 min, the cell mixture was subjected to low speed centrifugation and the supernatants, free of hemocytes, were collected. The number of C. albicans remaining in the supernatants was determined by colony counting. The data represent the mean ± SD of three experiments performed in triplicates.

Article Snippet: Galleria mellonella larvae inoculated with the wild-type Acinetobacter baumannii ATCC 17978 strain (1.1 × 10 5 CFU) died rapidly (100% death within 40 h) but not when inoculated with the Acinetobacter baumannii hdc::TOPO mutant (10% mortality).

Techniques: Produced, Mutagenesis, Concentration Assay, Enzyme-linked Immunosorbent Assay, Incubation, Centrifugation

Journal: Journal of Innate Immunity

Article Title: Histamine Produced by Gram-Negative Bacteria Impairs Neutrophil's Antimicrobial Response by Engaging the Histamine 2 Receptor

doi: 10.1159/000525536

Figure Lengend Snippet: Construction of the hdc-deficient A. baumannii ATCC 17978 strain. a A region of the hdc gene (homology region) was amplified by PCR using the genomic DNA of A. baumannii ATCC 17978 as a template. The PCR product was purified, and inserted in to the pCR-Blunt II-TOPO plasmid. Competent A. baumannii ATCC 17978 were electroporated with the pCR-Blunt II-TOPO plasmid containing the homology region of the hdc gene. Recombinants were selected on kanamycin plates. The predicted organization of the genomic DNA of the hdc recombinant hdc::TOPO is shown. To confirm that the hdc gene has been disrupted, the region between the T7 promoter (plasmid sequence) and a region upstream of the hdc gene were amplified by PCR using genomic DNA of the hdc::TOPO recombinant as a template. Two reverse primers (P1 rev or P2 rev) recognizing a gene sequence upstream of the hdc gene were designed. The predicted sizes of the amplicons are 843 and 844 bp with P1 rev and T7 forward primers and P2 rev and T7 forward primers, respectively. b The amplicons obtained by PCR using either genomic DNA from the recombinant hdc::TOPO or the WT strain as templates were separated on 1% agarose gel and visualized. The size of the molecular standards (in bp) is indicated on the left hand side of the gel. c the sequence of the amplicon obtained by PCR using the P1 rev and T7 forward primers and genomic DNA of the hdc mutant hdc::TOPO as a template is indicated. The gray part is the plasmid sequence upstream of the T7 promoter, followed by the sequence of the homology region, then the sequence of the end part of the hdc gene (underlined), and finally the genomic sequence 3′ upstream of the hdc gene (bold).

Article Snippet: Galleria mellonella larvae inoculated with the wild-type Acinetobacter baumannii ATCC 17978 strain (1.1 × 10 5 CFU) died rapidly (100% death within 40 h) but not when inoculated with the Acinetobacter baumannii hdc::TOPO mutant (10% mortality).

Techniques: Amplification, Purification, Plasmid Preparation, Recombinant, Sequencing, Agarose Gel Electrophoresis, Mutagenesis

Journal: Journal of Innate Immunity

Article Title: Histamine Produced by Gram-Negative Bacteria Impairs Neutrophil's Antimicrobial Response by Engaging the Histamine 2 Receptor

doi: 10.1159/000525536

Figure Lengend Snippet: WT A. baumannii ATCC 17978, but not hdc::TOPO mutant, produces histamine. a A. baumannii WT or hdc::TOPO mutant strains were inoculated in LB medium and grown in a glass flask under orbital rotation at 37°C in the absence or presence of histidine (10 −3 M). After 18 h, aliquots of bacteria were collected and spun down (2,500 g , 10 min). The resulting supernatants were transferred to 1.5 mL Eppendorf tubes and aliquots were diluted, and then used for the quantification of histamine by ELISA using the protocol provided by the manufacturer. The data are expressed as mean values ± SD of triplicates. b A culture of overnight grown A. baumannii ATCC 17978 or hdc::TOPO mutant were diluted in LB broth and adjusted to an OD value of 0.2 at 600 nm. The samples were then diluted 1/100 in LB medium and 10 mL of each mixture was transferred to 250 mL flasks. The flasks were put in an orbital shaker at 37°C. After each hour, an aliquot is taken out from the flask, diluted in LB medium, and turbidimetry at 600 nm was read. The OD values versus time are plotted. Wild type, dashed line; hdc::TOPO mutant, solid line.

Article Snippet: Galleria mellonella larvae inoculated with the wild-type Acinetobacter baumannii ATCC 17978 strain (1.1 × 10 5 CFU) died rapidly (100% death within 40 h) but not when inoculated with the Acinetobacter baumannii hdc::TOPO mutant (10% mortality).

Techniques: Mutagenesis, Bacteria, Enzyme-linked Immunosorbent Assay