Uncovering Differences in Virulence Markers Associated with Achromobacter Species of CF and Non-CF Origin

Achromobacter spp. are recognized as emerging pathogens in hospitalized as well as in cystic fibrosis (CF) patients. From 2012 to 2015, we collected 69 clinical isolates (41 patient) of Achromobacter spp. from 13 patients with CF (CF isolates, n = 32) and 28 patients receiving care for other health conditions (non-CF isolates, n = 37). Molecular epidemiology and virulence potential of isolates were examined. Antimicrobial susceptibility, motility, ability to form biofilms and binding affinity to mucin, collagen, and fibronectin were tested to assess their virulence traits. The nrdA gene sequencing showed that A. xylosoxidans was the most prevalent species in both CF and non-CF patients. CF patients were also colonized with A. dolens/A, ruhlandii, A. insuavis, and A. spiritinus strains while non-CF group was somewhat less heterogenous, although A. insuavis, A. insolitus, and A. piechaudii strains were detected beside A. xylosoxidans. Three strains displayed clonal distribution, one among patients from the CF group and two among non-CF patients. No significant differences in susceptibility to antimicrobials were observed between CF and non-CF patients. About one third of the isolates were classified as strong biofilm producers, and the proportion of CF and non-CF isolates with the ability to form biofilm was almost identical. CF isolates were less motile compared to the non-CF group and no correlation was found between swimming phenotype and biofilm formation. On the other hand, CF isolates exhibited higher affinity to bind mucin, collagen, and fibronectin. In generall, CF isolates from our study exhibited in vitro properties that could be of importance for the colonization of CF patients

Brevibacillus laterosporus strains BGSP7, BGSP9 and BGSP11 isolated from silage produce broad spectrum multi-antimicrobials

Bacteria active against multi-drug resistant pathogens, isolated by direct selection of colonies from clover silage samples, produce zones of inhibition against two Gram-negative (Klebsiella pneumoniae Ni9 and Pseudomonas aeruginosa MMA83) and two Gram-positive (Staphylococcus aureus ATCC25923 and Listeria monocytogenes ATCC19111) pathogens. Isolates BGSP7, BGSP9, BGSP11 and BGSP12 produced the largest zones of inhibition against all four pathogens when grown in LB broth with aeration at 37 degrees C. Isolates BGSP7, BGSP9, BGSP11 and BGSP12 were identified as Brevibacillus laterosporus and pulsed field gel electrophoresis and extracellular protein profiles showed that three different strains (BGSP7, BGSP9 and BGSP11) were isolated. A semi-native SDS-PAGE (sodium dodecyl sulphate-polyacrylamide gel electrophoresis) gel overlay assay showed that BGSP7 and BGSP9 produce small antimicrobial molecules of about 1.5 kDa, while BGSP11 produces antimicrobial molecules of 1.5 and 6 kDa active against S. aureus ATCC25923. Amino acid analysis of two antimicrobial molecules (1583.73 Da; from BGSP7 and 1556.31 Da; from BGSP11) revealed that they have a similar composition and differ only by virtue of the presence of a methionine which is present only in BGSP11 molecule. Genome sequencing of the three isolates revealed the presence of gene clusters associated with the production of non-ribosomally synthesized peptides (brevibacillin, bogorol, gramicidin S, plipastatin and tyrocin) and bacteriocins (laterosporulin, a lactococcin 972-like bacteriocin, as well as putative linocin M18, sactipeptide, UviB and lantipeptide-like molecules). Ultimately, the purification of a number of antimicrobial molecules from each isolate suggests that they can be considered as potent biocontrol strains that produce an arsenal of antimicrobial molecules active against Gram-positive and Gram-negative multi-resistant pathogens, fungi and insects

IC50 values, genotyping results and CarO variants for <i>A</i>. <i>baumannii</i> isolates from Serbia.

<p>IC50 values, genotyping results and CarO variants for <i>A</i>. <i>baumannii</i> isolates from Serbia.</p

Representation of β-lactamases detected in carbapenem-resistant <i>A</i>. <i>baumannii</i> clinical isolates from Serbia.

<p>Representation of β-lactamases detected in carbapenem-resistant <i>A</i>. <i>baumannii</i> clinical isolates from Serbia.</p

Dendrogram derived from <i>Apa</i>I PFGE patterns showing the relatedness of <i>A</i>. <i>baumannii</i> isolated in Serbia.

<p>The dendrogram was constructed using SPSS software. Letters A, B, C and D indicate different pulsotypes, while I and II designate two major clusters.</p

Phylogenetic inferences of CarO protein among <i>Acinetobacter</i> spp.

<p>A phylogenetic tree of CarO proteins was constructed with the maximum likelihood (ML) method using a Jones-Taylor-Thornton (JTT) model distance matrix. The confidence levels were calculated from 1000 bootstrap resamples of alignment used for phylogenetic inferences by ML method. Bold gray and bold black lines represent the nodes with a support bootstrap value of ≥50% and ≥70%, respectively. The black triangles represent the clade consisting of only <i>A</i>. <i>baumannii</i> strains from the database. Gene bank accession numbers for all tree members are given in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0122793#pone.0122793.s001" target="_blank">S1 Fig</a> Representatives and variants (I-VI) of each group (I-III) are the following: <i>A</i>. <i>baumannii</i> Ab21 (DQ309875), <i>A</i>. <i>baumannii</i> Ab244 (AY684798), <i>A</i>. <i>baumannii</i> Ab413 (FJ652395), <i>A</i>. <i>baumannii</i> Ab253 (EF537047), <i>A</i>. <i>baumannii</i> 146457 (EXB49165), <i>A</i>. <i>baumannii</i> 230853 (EXB72592), <i>A</i>. <i>baumannii</i> 1461402 (EXB34375), <i>A</i>. <i>baumannii</i> 348935 (EXA64785). The Serbian clinical isolates <i>A</i>. <i>baumannii</i> 6000 and <i>A</i>. <i>baumannii</i> 1955/12 are indicated.</p

Primers used in this study.

<p>Primers used in this study.</p