A New Method for Determination of Minimum Biofilm Eradication Concentration for Accurate Antimicrobial Therapy

Ravi, Nithin Sam; Aslam, Raziya Fathima; Veeraraghavan, Balaji

doi:10.1007/978-1-4939-9118-1_6

Cited by 13 publications

(10 citation statements)

References 5 publications

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“…Nevertheless, some cells remained viable in the biofilm and proliferate when transferring suspension onto agar plates. Therefore, unlike Ravi et al [ 45 ], in our study, use of the resazurin colorimetric assay alone did not allow us to obtain reliable MBEC values in existing biofilms. These data suggested that some biofilm cells remain dormant due to cell stress, preventing the reduction of resazurin.…”

Section: Discussioncontrasting

confidence: 72%

“…The plates were incubated at 37 °C overnight. The minimum fungicidal concentration (MFC) is defined as the lowest concentration of compound that yielded no Candida growth on agar plates, and the minimum biofilm eliminating concentration (MBEC) is defined as the lowest concentration resulting in a 100% cells death of the biofilm (no growth observed on agar plates) [ 45 , 46 ].…”

Section: Methodsmentioning

confidence: 99%

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In Vitro Activity of Carbosilane Cationic Dendritic Molecules on Prevention and Treatment of Candida Albicans Biofilms

et al. 2020

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Candida spp. are one of the most common fungal pathogens. Biofilms formed by Candidaalbicans offer resistance mechanisms against most antifungal agents. Therefore, development of new molecules effective against these microorganisms, alone or in combination with antifungal drugs, is extremely necessary. In the present work, we carried out a screening process of different cationic carbosilane dendritic molecules against C. albicans. In vitro activity against biofilm formation and biofilms was tested in both Colección Española de Cultivos Tipo (CECT) 1002 and clinical C. albicans strains. Cytotoxicity was studied in human cell lines, and biofilm alterations were observed by scanning electron microscopy (SEM). Antifungal activity of the carbosilane dendritic molecules was assessed by monitoring cell viability using both established and novel cell viability assays. One out of 14 dendritic molecules tested, named BDSQ024, showed the highest activity with a minimum biofilm inhibitory concentration (MBIC) for biofilm formation and a minimum biofilm damaging concentration (MBDC) for existing biofilm of 16–32 and 16 mg/L, respectively. Synergy with amphotericin (AmB) and caspofungin (CSF) at non-cytotoxic concentrations was found. Therefore, dendritic compounds are exciting new antifungals effective at preventing Candida biofilm formation and represent a potential novel therapeutic agent for treatment of C. albicans infection in combination with existing clinical antifungals.

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Section: Discussioncontrasting

confidence: 72%

Section: Methodsmentioning

confidence: 99%

In Vitro Activity of Carbosilane Cationic Dendritic Molecules on Prevention and Treatment of Candida Albicans Biofilms

et al. 2020

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“…Furthermore, to assess the efficacy of the dispersal treatment, Gram staining of treated and untreated samples was performed. Simultaneously, a second quantification was performed by means of the resazurin assay, as described by Ravi et al (2019). Briefly, a standard plate containing 1-fold difference of a known cellular concentration between rows and 10-fold difference between columns was prepared from an overnight culture in a 96-well microplate for the fluorescence-based assays (Invitrogen).…”

Section: Antibiotic Treatment Exposurementioning

confidence: 99%

Identification and Characterization of Planktonic Biofilm-Like Aggregates in Infected Synovial Fluids From Joint Infections

et al. 2020

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Recent in vitro studies reported the exceptional ability of some bacterial species to form biofilm-like aggregates in human and animal synovial fluids (SF), but evidences from infected clinical samples are still lacking. In this study, we investigated whether this bacterial phenotype was present in infected SFs collected from joint infections and if it was maintained in in vitro settings. SFs sent for culture to the Laboratory of Microbiology of our institute were directly analyzed by means of confocal laser scanning microscopy (CLSM), and the infective agents were isolated for further in vitro tests. Moreover, sterile SF was collected from patients who did not receive previous antibiotic therapy to investigate the formation of bacterial aggregates, together with biofilm and matrix production on a titanium surface. Finally, antibiotic susceptibility studies were performed by using bovine SF. Four Staphylococcus aureus, one Staphylococcus lugdunensis, and one Prevotella bivia strain were identified in the infected SFs. The CLSM analysis showed that all staphylococci were present as a mixture of single cells and bacterial clumps surrounded by an exopolymeric substance, which comprised SF-derived fibrin, while all P. bivia cells appeared separated. Despite that, differences in the ability to aggregate between S. aureus and S. lugdunensis were observed in clinical SFs. These different phenotypes were further confirmed by in vitro growth, even though the application of such ex vivo approach lead all staphylococci to form exceptionally large microbial aggregates, which are several folds bigger than those observed in clinical samples. Planktonic aggregates challenged for antibiotic susceptibility revealed a sharp increase of recalcitrance to the treatments. Although this is still at a preliminary stage, the present work confirmed the ability of staphylococci to form free-floating biofilm-like aggregates in infected SF from patients with joint infections. Furthermore, the obtained results pointed out that future in vitro research on joint infections will benefit from the use of human-or animal-derived SF. Even though this approach should be carefully validated in further studies comprising a larger microbial population, these findings pose new challenges in the treatment of infected native and prosthetic joints and for the approach to new investigations.

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“…The minimum bacterial concentration (MBC) value for the pre-treatment assay, the lowest concentration that eliminated 100% of the cells stimulated to generate a biofilm, and the minimum biofilm eliminating concentration (MBEC) value for the post-biofilm treatment assay, the lowest concentration capable of completely eradicating bacteria from the previously established biofilm, were obtained by the drop plate method observing no growth on agar plates [ 28 , 29 , 30 , 31 ]. To perform these experiments, wells were scraped with the pipette tips to detach the biofilm cells and 5 μL of each well suspension were transferred to PCA agar plates and incubated overnight at 37 °C.…”

Section: Methodsmentioning

confidence: 99%

Effect of the Combination of Levofloxacin with Cationic Carbosilane Dendron and Peptide in the Prevention and Treatment of Staphylococcus aureus Biofilms

et al. 2021

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Antibiotic resistance and biofilm-related infections, persistent in conventional antimicrobial treatment, are continuously increasing and represent a major health problem worldwide. Therefore, the development of new effective treatments to prevent and treat biofilm-related infections represents a crucial challenge. Unfortunately, the extensive use of antibiotics has led to an increase of resistant bacteria with the subsequent loss of effectivity of commercial antibiotics, mainly due to antibiotic resistance and the ability of some bacteria to form microbial communities in biotic or abiotic surfaces (biofilms). In some cases, these biofilms are resistant to high concentrations of antibiotics that lead to treatment failure and recurrence of the associated infections. In the fight against microbial resistance, the combination of traditional antibiotics with new compounds (combination therapy) is an alternative that is becoming more extensive in the medical field. In this work, we studied the cooperative effects between levofloxacin, an approved antibiotic, and peptides or cationic dendritic molecules, compounds that are emerging as a feasible solution to overcome the problem of microbial resistance caused by pathogenic biofilms. We studied a new therapeutic approach that involves the use of levofloxacin in combination with a cationic carbosilane dendron, called MalG2(SNHMe2Cl)4, or a synthetic cell-penetrating peptide, called gH625, conjugated to the aforementioned dendron. To carry out the study, we used two combinations (1) levofloxacin/dendron and (2) levofloxacin/dendron-peptide nanoconjugate. The results showed the synergistic effect of the combination therapy to treat Staphylococcus aureus biofilms. In addition, we generated a fluorescein labeled peptide that allowed us to observe the conjugate (dendron-peptide) localization throughout the bacterial biofilm by confocal laser scanning microscopy.

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A New Method for Determination of Minimum Biofilm Eradication Concentration for Accurate Antimicrobial Therapy

Cited by 13 publications

References 5 publications

In Vitro Activity of Carbosilane Cationic Dendritic Molecules on Prevention and Treatment of Candida Albicans Biofilms

In Vitro Activity of Carbosilane Cationic Dendritic Molecules on Prevention and Treatment of Candida Albicans Biofilms

Identification and Characterization of Planktonic Biofilm-Like Aggregates in Infected Synovial Fluids From Joint Infections

Effect of the Combination of Levofloxacin with Cationic Carbosilane Dendron and Peptide in the Prevention and Treatment of Staphylococcus aureus Biofilms

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