Phenotypic and genotypic characterization of Staphylococci causing breast peri-implant infections in oncologic patients

Background

Staphylococcus epidermidis and S. aureus have been identified as the most common bacteria responsible for sub-clinical and overt breast implant infections and their ability to form biofilm on the implant as been reported as the essential factor in the development of this type of infections. Biofilm formation is a complex process with the participation of several distinct molecules, whose relative importance in different clinical settings has not yet been fully elucidated. To our knowledge this is the first study aimed at characterizing isolates causing breast peri-implant infections.

Results

Thirteen S. aureus and seven S. epidermidis causing breast peri-implant infections were studied.Using the broth microdilution method and the E-test, the majority of the strains were susceptible to all antibiotics tested. Methicillin resistance was detected in two S. epidermidis. All strains had different RAPD profiles and were able to produce biofilms in microtitre plate assays but, while all S. aureus carried and were able to express icaA and icaD genes, this was only true for one S. epidermidis. Biofilm development was glucose- and NaCl-induced (5 S. aureus and 1 S. epidermidis) or glucose-induced (the remaining strains). Proteinase K and sodium metaperiodate treatment had different effects on biofilms dispersion revealing that the strains studied were able to produce chemically different types of extracellular matrix mediating biofilm formation.All S. aureus strains harboured and expressed the atlA, clfA, FnA, eno and cna genes and the majority also carried and expressed the sasG (10/13), ebpS (10/13) genes.All S. epidermidis strains harboured and expressed the atlE, aae, embp genes, and the majority (six strains) also carried and expressed the fbe, aap genes.Genes for S. aureus capsular types 5 and 8 were almost equally distributed. The only leukotoxin genes detected were lukE/lukD (6/13).

Conclusions

S. aureus and S. epidermidis breast peri-implant infections are caused by heterogeneous strains with different biofilm development mechanisms.Since the collagen adhesin (cna) gene is not ubiquitously distributed among S. aureus, this protein could have an important role in the cause of breast peri-implant infections.

Electronic supplementary material

The online version of this article (doi:10.1186/s12866-015-0368-x) contains supplementary material, which is available to authorized users.     

Crystal structure of the functional region of Uro-adherence factor A from Staphylococcus saprophyticus reveals participation of the B domain in ligand binding

Staphylococci use cell wall‐anchored proteins as adhesins to attach to host tissues. Staphylococcus saprophyticus, a uropathogenic species, has a unique cell wall‐anchored protein, uro‐adherence factor A (UafA), which shows erythrocyte binding activity. To investigate the mechanism of adhesion by UafA, we determined the crystal structure of the functional region of UafA at 1.5 Å resolution. The structure was composed of three domains, designated as the N2, N3, and B domains, arranged in a triangular relative configuration. Hemagglutination inhibition assay with domain‐truncated mutants indicated that both N and B domains were necessary for erythrocyte binding. Based on these results, a novel manner of ligand binding in which the B domain acts as a functional domain was proposed as the adhesion mechanism of S. saprophyticus.     

Pneumococcal Adhesins PavB and PspC Are Important for the Interplay with Human Thrombospondin-1

The human matricellular glycoprotein thrombospondin-1 (hTSP-1) is released by activated platelets and mediates adhesion of Gram-positive bacteria to various host cells. In staphylococci, the adhesins extracellular adherence protein (Eap) and autolysin (Atl), both surface-exposed proteins containing repeating structures, were shown to be involved in the acquisition of hTSP-1 to the bacterial surface. The interaction partner(s) on the pneumococcal surface was hitherto unknown. Here, we demonstrate for the first time that pneumococcal adherence and virulence factor B (PavB) and pneumococcal surface protein C (PspC) are key players for the interaction of Streptococcus pneumoniae with matricellular hTSP-1. PavB and PspC are pneumococcal surface-exposed adhesins and virulence factors exhibiting repetitive sequences in their core structure. Heterologously expressed fragments of PavB and PspC containing repetitive structures exhibit hTSP-1 binding activity as shown by ELISA and surface plasmon resonance studies. Binding of hTSP-1 is charge-dependent and inhibited by heparin. Importantly, the deficiency in PavB and PspC reduces the recruitment of soluble hTSP-1 by pneumococci and decreases hTSP-1-mediated pneumococcal adherence to human epithelial cells. Platelet activation assays suggested that PavB and PspC are not involved in the activation of purified human platelets by pneumococci. In conclusion, this study indicates a pivotal role of PavB and PspC for pneumococcal recruitment of soluble hTSP-1 to the bacterial surface and binding of pneumococci to host cell-bound hTSP-1 during adhesion.     

Pyrrolomycins as antimicrobial agents. Microwave-assisted organic synthesis and insights into their antimicrobial mechanism of action

New compounds able to counteract staphylococcal biofilm formation are needed. In this study we investigate the mechanism of action of pyrrolomycins, whose potential as antimicrobial agents has been demonstrated. We performed a new efficient and easy method to use microwave organic synthesis suitable for obtaining pyrrolomycins in good yields and in suitable amount for their in vitro in-depth investigation. We evaluate the inhibitory activity towards Sortase A (SrtA), a transpeptidase responsible for covalent anchoring in Gram-positive peptidoglycan of many surface proteins involved in adhesion and in biofilm formation. All compounds show a good inhibitory activity toward SrtA, having IC₅₀ values ranging from 130 to 300?µM comparable to berberine hydrochloride. Of note compound 1d shows a good affinity in docking experiment to SrtA and exhibits the highest capability to interfere with biofilm formation of S. aureus showing an IC₅₀ of 3.4?nM. This compound is also effective in altering S. aureus murein hydrolase activity that is known to be responsible for degradation, turnover, and maturation of bacterial peptidoglycan and involved in the initial stages of S. aureus biofilm formation.     

Bacterial virulence as a target for antimicrobial chemotherapy

As bacterial resistance to currently used antibiotics increases, so too must efforts to identify novel agents and strategies for the prevention and treatment of bacterial infection. In the past, antimicrobial drug discovery efforts have focused on eradicating infection by either cidal or static agents, resulting in clearance of the bacterium from the infected host. To this end, drug discovery targets have been those proteins or processes essential for bacterial cell viability. However, inhibition of the interaction between the bacterium and its host may also be a target. During establishment of an infection, pathogenic bacteria use carefully regulated pathways of conditional gene expression to transition from a free-living form to one that must adapt to the host milieu. This transition requires the regulated production of both extracellular and cell-surface molecules, often termed virulence factors. As the biological imperatives of the invading organism change during the course of an infection, the expression of these factors is altered in response to environmental cues. These may be changes in the host environment, for example, pH, metabolites, metal ions, osmolarity, and temperature. Alternatively, effector molecules produced by the bacterium to sense changing cell density can also lead to changes in virulence gene expression. Although the mechanisms of pathogenesis among different bacteria vary, the principles of virulence are generally conserved. Bacterial virulence may therefore offer unique opportunities to inhibit the establishment of infection or alter its course as a method of antimicrobial chemotherapy.