Microbial biofilm: a “sticky” problem

Bacteria can form, on virtually any surface, single- and multispecies biofilms intrinsically resistant/tolerant to antibiotics and elusive of the host immune response. The study of bacterial biofilm development has, therefore, received great interest over the past 20 years and is motivated by the well-recognized role of these multicellular communities in infectious diseases. In this review article, we provide a synopsis of (i) biofilm formation mechanisms; (ii) biofilm clinical significance and underlying mechanisms; (iii) the current methodologies for microbiological diagnosis of biofilm-related infections; and (iv) current and future therapeutic strategies to combat biofilm-associated infections

D-BMAP18 antimicrobial peptide is active In Vitro, resists to pulmonary proteases but loses its activity in a murine model of Pseudomonas aeruginosa lung infection

The spread of antibiotic resistant-pathogens is driving the search for new antimicrobial compounds. Pulmonary infections experienced by cystic fibrosis (CF) patients are a dramatic example of this health-care emergency. Antimicrobial peptides could answer the need for new antibiotics but translating them from basic research to the clinic is a challenge. We have previously evaluated the potential of the small membranolytic peptide BMAP-18 to treat CF-related infections, discovering that while this molecule had a good activity in vitro it was not active in vivo because of its rapid degradation by pulmonary proteases. In this study, we synthesized and tested the proteases-resistant all-d enantiomer. In spite of a good antimicrobial activity against Pseudomonas aeruginosa and Stenotrophomonas maltophilia clinical isolates and of a tolerable cytotoxicity in vitro, D-BMAP18 was ineffective to treat P. aeruginosa pulmonary infection in mice, in comparison to tobramycin. We observed that different factors other than peptide degradation hampered its efficacy for pulmonary application. These results indicate that D-BMAP18 needs further optimization before being suitable for clinical application and this approach may represent a guide for optimization of other anti-infective peptides eligible for the treatment of pulmonary infections

Stenotrophomonas maltophilia Phenotypic and Genotypic Diversity during a 10-year Colonization in the Lungs of a Cystic Fibrosis Patient

The present study was carried out to understand the adaptive strategies developed by Stenotrophomonas maltophilia for chronic colonization of the cystic fibrosis (CF) lung. For this purpose, 13 temporally isolated strains from a single CF patient chronically infected over a 10-year period were systematically characterized for growth rate, biofilm formation, motility, mutation frequencies, antibiotic resistance, and pathogenicity. Pulsed-field gel electrophoresis (PFGE) showed over time the presence of two distinct groups, each consisting of two different pulsotypes. The pattern of evolution followed by S. maltophilia was dependent on pulsotype considered, with strains belonging to pulsotype 1.1 resulting to be the most adapted, being significantly changed in all traits considered. Generally, S. maltophilia adaptation to CF lung leads to increased growth rate and antibiotic resistance, whereas both in vivo and in vitro pathogenicity as well as biofilm formation were decreased. Overall, our results show for the first time that S. maltophilia can successfully adapt to a highly stressful environment such as CF lung by paying a "biological cost," as suggested by the presence of relevant genotypic and phenotypic heterogeneity within bacterial population. S. maltophilia populations are, therefore, significantly complex and dynamic being able to fluctuate rapidly under changing selective pressures

In vitro and in vivo evaluation of BMAP-derived peptides for the treatment of cystic fibrosis-related pulmonary infections

Patients with cystic fibrosis require pharmacological treatment against chronic lung infections. The alpha-helical antimicrobial peptides BMAP-27 and BMAP-28 have shown to be highly active in vitro against planktonic and sessile forms of multidrug-resistant Pseudomonas aeruginosa, Staphylococcus aureus, and Stenotrophomonas maltophilia cystic fibrosis strains. To develop small antibacterial peptides for therapeutic use, we tested shortened/modified BMAP fragments, and selected the one with the highest in vitro antibacterial activity and lowest in vivo acute pulmonary toxicity. All the new peptides have shown to roughly maintain their antibacterial activity in vitro. The 1-18 N-terminal fragment of BMAP-27, showing MIC90 of 16 A mu g/ml against P. aeruginosa isolates and strain-dependent anti-biofilm effects, showed the lowest pulmonary toxicity in mice. However, when tested in a murine model of acute lung infection by P. aeruginosa, BMAP-27(1-18) did not show any curative effect. If exposed to murine broncho-alveolar lavage fluid BMAP-27(1-18) was degraded within 10 min, suggesting it is not stable in pulmonary environment, probably due to murine proteases. Our results indicate that shortened BMAP peptides could represent a starting point for antibacterial drugs, but they also indicate that they need a further optimization for effective in vivo use

Adhesion to and biofilm formation on IB3-1 bronchial cells by Stenotrophomonas maltophilia isolates from cystic fibrosis patients

<p>Abstract</p> <p>Background</p> <p><it>Stenotrophomonas maltophilia </it>has recently gained considerable attention as an important emerging pathogen in cystic fibrosis (CF) patients. However, the role of this microorganism in the pathophysiology of CF lung disease remains largely unexplored. In the present study for the first time we assessed the ability of <it>S. maltophilia </it>CF isolates to adhere to and form biofilm in experimental infection experiments using the CF-derived bronchial epithelial IB3-1cell line. The role of flagella on the adhesiveness of <it>S. maltophilia </it>to IB3-1 cell monolayers was also assessed by using <it>fliI </it>mutant derivative strains.</p> <p>Results</p> <p>All <it>S. maltophilia </it>CF isolates tested in the present study were able, although at different levels, to adhere to and form biofilm on IB3-1 cell monolayers. Scanning electron and confocal microscopy revealed <it>S. maltophilia </it>structures typical of biofilm formation on bronchial IB3-1 cells. The loss of flagella significantly (P < 0.001) decreased bacterial adhesiveness, if compared to that of their parental flagellated strains. <it>S. maltophilia </it>CF isolates were also able to invade IB3-1 cells, albeit at a very low level (internalization rate ranged from 0.01 to 4.94%). Pre-exposure of IB3-1 cells to <it>P. aeruginosa </it>PAO1 significantly increased <it>S. maltophilia </it>adhesiveness. Further, the presence of <it>S. maltophilia </it>negatively influenced <it>P. aeruginosa </it>PAO1 adhesiveness.</p> <p>Conclusions</p> <p>The main contribution of the present study is the finding that <it>S. maltophilia </it>is able to form biofilm on and invade CF-derived IB3-1 bronchial epithelial cells, thus posing a rationale for the persistence and the systemic spread of this opportunistic pathogen in CF patients. Experiments using <it>in vivo </it>models which more closely mimic CF pulmonary tissues will certainly be needed to validate the relevance of our results.</p

Phenotypic and genotypic characterization of Stenotrophomonas maltophilia isolates from patients with cystic fibrosis: Genome diversity, biofilm formation, and virulence

<p>Abstract</p> <p>Background</p> <p><it>Stenotrophomonas maltophilia </it>is emerging as one of the most frequently found bacteria in cystic fibrosis (CF) patients. In the present study, phenotypic and genotypic traits of a set of 98 isolates of <it>S. maltophilia </it>obtained from clinical (CF and non-CF patients) and environmental sources were comparatively evaluated.</p> <p>Results</p> <p><it>S. maltophilia </it>exhibited a high level of genomic diversity in both CF and non-CF group, thus possibly allowing this bacterium to expand its pathogenic potentials. Strains sharing the same pulsotype infected different patients, thus likely indicating the occurrence of clonal spread or acquisition by a common source. CF isolates differed greatly in some phenotypic traits among each other and also when compared with non-CF isolates, demonstrating increased mean generation time and susceptibility to oxidative stress, but reduced ability in forming biofilm. Furthermore, in CF isolates flagella- and type IV pili-based motilities were critical for biofilm development, although not required for its initiation. Sequential isogenic strains isolated from the same CF patient displayed heterogeneity in biofilm and other phenotypic traits during the course of chronic infection. CF and non-CF isolates showed comparable virulence in a mouse model of lung infection.</p> <p>Conclusions</p> <p>Overall, the phenotypic differences observed between CF and non-CF isolates may imply different selective conditions and persistence (adaptation) mechanisms in a hostile and heterogeneous environment such as CF lung. Molecular elucidation of these mechanisms will be essential to better understand the selective adaptation in CF airways in order to design improved strategies useful to counteract and eradicate <it>S. maltophilia </it>infection.</p

The anti-microbial peptide (Lin-SB056-1)(2)-K reduces pro-inflammatory cytokine release through Interaction with Pseudomonas aeruginosa lipopolysaccharide

The ability of many anti-microbial peptides (AMPs) to modulate the host immune response has highlighted their possible therapeutic use to reduce uncontrolled inflammation during chronic infections. In the present study, we examined the anti-inflammatory potential of the semi-synthetic peptide lin-SB056-1 and its dendrimeric derivative (lin-SB056-1)(2)-K, which were previously found to have anti-microbial activity against Pseudomonas aeruginosa in in vivo-like models mimicking the challenging environment of chronically infected lungs (i.e., artificial sputum medium and 3-D lung mucosa model). The dendrimeric derivative exerted a stronger anti-inflammatory activity than its monomeric counterpart towards lung epithelial- and macrophage-cell lines stimulated with P. aeruginosa lipopolysaccharide (LPS), based on a marked decrease (up to 80%) in the LPS-induced production of different pro-inflammatory cytokines (i.e., IL-1 beta, IL-6 and IL-8). Accordingly, (lin-SB056-1)(2)-K exhibited a stronger LPS-binding affinity than its monomeric counterpart, thereby suggesting a role of peptide/LPS neutralizing interactions in the observed anti-inflammatory effect. Along with the anti-bacterial and anti-biofilm properties, the anti-inflammatory activity of (lin-SB056-1)(2)-K broadens its therapeutic potential in the context of chronic (biofilm-associated) infections

Bioactive compounds: a goldmine for defining new strategies against pathogenic bacterial biofilms?

Most human infectious diseases are caused by microorganisms growing as biofilms. These three-dimensional self-organized communities are embedded in a dense matrix allowing microorganisms to persistently inhabit abiotic and biotic surfaces due to increased resistance to both antibiotics and effectors of the immune system. Consequently, there is an urgent need for novel strategies to control biofilm-associated infections. Natural products offer a vast array of chemical structures and possess a wide variety of biological properties; therefore, they have been and continue to be exploited in the search for potential biofilm inhibitors with a specific or multi-locus mechanism of action. This review provides an updated discussion of the major bioactive compounds isolated from several natural sources - such as plants, lichens, algae, microorganisms, animals, and humans - with the potential to inhibit biofilm formation and/or to disperse established biofilms by bacterial pathogens. Despite the very large number of bioactive products, their exact mechanism of action often remains to be clarified and, in some cases, the identity of the active molecule is still unknown. This knowledge gap should be filled thus allowing development of these products not only as novel drugs to combat bacterial biofilms, but also as antibiotic adjuvants to restore the therapeutic efficacy of current antibiotics

An Overview on <i>Streptococcus bovis/Streptococcus equinus</i> Complex Isolates: Identification to the Species/Subspecies Level and Antibiotic Resistance

Streptococcus bovis/Streptococcus equinus complex (SBSEC), a non-enterococcal group D Streptococcus spp. complex, has been described as commensal bacteria in humans and animals, with a fecal carriage rate in humans varying from 5% to over 60%. Among streptococci, SBSEC isolates represent the most antibiotic-resistant species&#8212;with variable resistance rates reported for clindamycin, erythromycin, tetracycline, and levofloxacin&#8212;and might act as a reservoir of multiple acquired genes. Moreover, reduced susceptibility to penicillin and vancomycin associated with mobile genetic elements have also been detected, although rarely. Since the association of SBSEC bacteremia and colon lesions, infective endocarditis and hepatobiliary diseases has been established, particularly in elderly individuals, an accurate identification of SBSEC isolates to the species and subspecies level, as well as the evaluation of antibiotic resistance, are needed. In this paper, we reviewed the major methods used to identify SBSEC isolates and the antimicrobial resistance rates reported in the scientific literature among SBSEC species