Prevalence of the Genes Associated with Biofilm and Toxins Synthesis amongst the Pseudomonas aeruginosa Clinical Strains

Pseudomonas aeruginosa is one of the most commonly isolated bacteria from clinical specimens, with an increasing isolation frequency in nosocomial outbreaks. The hypothesis tested was whether carbapenem-resistant P. aeruginosa strains display an altered carriage of the virulence factor genes, depending on the type of carbapenem resistance. The aim of the study was to investigate, by PCR, the frequency of 10 chosen virulence factors genes (phzM, phzS, exoT, exoY, exoU, toxA, exoS, algD, pilA and pilB) and the genotype distribution in 107 non-duplicated carbapenem-resistant P. aeruginosa isolates. P. aeruginosa genes involved in phenazine dyes and exoenzyme T synthesis were noted with the highest frequency (100%). Fimbriae-encoding genes were detected with the lowest incidence: 15.9% and 4.7% for pilin A and B, respectively. The differences observed between the exoS gene prevalence amongst the carbapenemase-positive and the carbapenemase-negative strains and the pilA gene prevalence amongst the strains of different origins were statistically significant. Virulence genes’ prevalence and the genotype distribution vary amongst P. aeruginosa strains resistant to carbapenems, especially in terms of their carbapenemase synthesis ability and the strain origin.

A rapid method for the detection of motility in Acinetobacter baumannii and its association to the existence of the virulence-associated genes pilA and algW

Acinetobacter (A.) baumannii is one of the major nosocomial pathogens worldwide. It is associated with bloodstream infection, pneumonia, meningitis, urinary tract, soft tissue, and wound infections. Several factors contribute to its survival and spread as a nosocomial pathogen, and motility is often associated with the virulence, fitness, and tenacity of A. baumannii on surfaces. In the present study, the correlation between the presence of genes encoding for fimbrial protein PilA and periplasmic protease AlgW and motility was investigated in 87 clinical and non-clinical A. baumannii isolates from Germany. A. baumannii exhibited robust swimming, swarming, and twitching movement based on the percentage of agar in the medium, as well as the time and temperature of incubation. The swarm motility medium utilizing 2% agar with tetrazolium salts provided an efficient assay for the phenotypic characterization of A. baumannii and it was more efficient than the classical motility assays in terms of time, visibility, and biosafety. The presence of the pilA gene increased motility of A. baumannii but was not required for motility. The algW gene was found in 18 strains obtained from milk, all of them with proven phenotypic motility. The rapid detection of motility is essential to evaluate the virulence and fitness of A. baumannii. Further studies on the level of genome, transcriptome and proteome are needed to investigate the secrets behind different movement paths in each strain.

Characterization of Type IV Pilus Genes in Pseudomonas syringae pv. tomato DC3000

Many strains of Pseudomonas syringae produce retractile pili that act as receptors for lytic bacteriophage φ6. As these are also characteristics of type IV pili, it was postulated that P. syringae may possess genes for type IV pilus biogenesis. A cosmid clone bank of P. syringae pv. tomato DC3000 genomic DNA was used to complement a mutant of Pseudomonas aeruginosa defective in the PilD (XcpA) prepilin peptidase gene by selection for restoration of extracellular protein secretion, a function also known to require PilD. A cosmid able to complement this mutant was also able to complement mutations in the pilB and pilC genes, suggesting that, if the organization of these genes is similar to that of P. aeruginosa, the cosmid may contain the P. syringae pilA. This was confirmed by sequencing a region from this plasmid that was shown to hybridize at low stringency to the P. aeruginosa pilA gene. The deduced P. syringae PilA polypeptide possesses the characteristic properties of the type IV pilins. Heterologous expression of the P. syringae pilA in P. aeruginosa was also shown, conferring not only φ6 phage sensitivity to P. aeruginosa pilA mutants but also sensitivity to PO4, a lytic bacteriophage specific for the pilus of P. aeruginosa. This suggests that additional components might be present in the mature pilus of P. aeruginosa that are the true receptors for this phage. Chromosomal mutations in P. syringae pv. tomato DC3000 pilA and pilD genes were shown to abolish its sensitivity to bacteriophage φ6. To determine the importance of P. syringae pilus in plant leaf interactions, these mutations were tested under laboratory and field conditions. Although little effect was seen on pathogenicity, culturable leaf-associated population sizes of the pilA mutant were significantly different from those of the wild-type parent. In addition, the expression of the DC3000 pilA gene appears to contribute to the UV tolerance of P. syringae and may play a role in survival on the plant leaf surface.