skip to main content

Title: Antibiotic Susceptibility of Escherichia coli Cells during Early-Stage Biofilm Formation
ABSTRACT Bacteria form complex multicellular structures on solid surfaces known as biofilms, which allow them to survive in harsh environments. A hallmark characteristic of mature biofilms is the high-level antibiotic tolerance (up to 1,000 times) compared with that of planktonic cells. Here, we report our new findings that biofilm cells are not always more tolerant to antibiotics than planktonic cells in the same culture. Specifically, Escherichia coli RP437 exhibited a dynamic change in antibiotic susceptibility during its early-stage biofilm formation. This phenomenon was not strain specific. Upon initial attachment, surface-associated cells became more sensitive to antibiotics than planktonic cells. By controlling the cell adhesion and cluster size using patterned E. coli biofilms, cells involved in the interaction between cell clusters during microcolony formation were found to be more susceptible to ampicillin than cells within clusters, suggesting a role of cell-cell interactions in biofilm-associated antibiotic tolerance. After this stage, biofilm cells became less susceptible to ampicillin and ofloxacin than planktonic cells. However, when the cells were detached by sonication, both antibiotics were more effective in killing the detached biofilm cells than the planktonic cells. Collectively, these results indicate that biofilm formation involves active cellular activities in adaption to the attached life more » form and interactions between cell clusters to build the complex structure of a biofilm, which can render these cells more susceptible to antibiotics. These findings shed new light on bacterial antibiotic susceptibility during biofilm formation and can guide the design of better antifouling surfaces, e.g., those with micron-scale topographic structures to interrupt cell-cell interactions. IMPORTANCE Mature biofilms are known for their high-level tolerance to antibiotics; however, antibiotic susceptibility of sessile cells during early-stage biofilm formation is not well understood. In this study, we aim to fill this knowledge gap by following bacterial antibiotic susceptibility during early-stage biofilm formation. We found that the attached cells have a dynamic change in antibiotic susceptibility, and during certain phases, they can be more sensitive to antibiotics than planktonic counterparts in the same culture. Using surface chemistry-controlled patterned biofilm formation, cell-surface and cell-cell interactions were found to affect the antibiotic susceptibility of attached cells. Collectively, these findings provide new insights into biofilm physiology and reveal how adaptation to the attached life form may influence antibiotic susceptibility of bacterial cells. « less
Authors:
; ; ; ; ;
Award ID(s):
1706061
Publication Date:
NSF-PAR ID:
10162427
Journal Name:
Journal of Bacteriology
Volume:
201
Issue:
18
ISSN:
0021-9193
Sponsoring Org:
National Science Foundation
More Like this
  1. The rise in bacterial resistance to common antibiotics has raised an increased need for alternative treatment strategies. The natural antibacterial product, 18β-glycyrrhetinic acid (GRA) has shown efficacy against community-associated methicillin-resistant Staphylococcus aureus (MRSA), although its interactions against planktonic and biofilm modes of growth remain poorly understood. This investigation utilized biochemical and metabolic approaches to further elucidate the effects of GRA on MRSA. Prolonged exposure of planktonic MRSA cell cultures to GRA resulted in increased production of staphyloxanthin, a pigment known to exhibit antioxidant and membrane-stabilizing functions. Then, 1D 1H NMR analyses of intracellular metabolite extracts from MRSA treated with GRA revealed significant changes in intracellular polar metabolite profiles, including increased levels of succinate and citrate, and significant reductions in several amino acids, including branch chain amino acids. These changes reflect the MRSA response to GRA exposure, including potentially altering its membrane composition, which consumes branched chain amino acids and leads to significant energy expenditure. Although GRA itself had no significant effect of biofilm viability, it seems to be an effective biofilm disruptor. This may be related to interference with cell–cell aggregation, as treatment of planktonic MRSA cultures with GRA leads to a significant reduction in micro-aggregation. The dispersive nature ofmore »GRA on MRSA biofilms may prove valuable for treatment of such infections and could be used to increase susceptibility to complementary antibiotic therapeutics.« less
  2. Many bacterial species in nature possess the ability to transition into a sessile lifestyle and aggregate into cohesive colonies, known as biofilms. Within a biofilm, bacterial cells are encapsulated within an extracellular polymeric substance (EPS) comprised of polysaccharides, proteins, nucleic acids, lipids, and other small molecules. The transition from planktonic growth to the biofilm lifecycle provides numerous benefits to bacteria, such as facilitating adherence to abiotic surfaces, evasion of a host immune system, and resistance to common antibiotics. As a result, biofilm-forming bacteria contribute to 65% of infections in humans, and substantially increase the energy and time required for treatment and recovery. Several biofilm specific exopolysaccharides, including cellulose, alginate, Pel polysaccharide, and poly- N -acetylglucosamine (PNAG), have been shown to play an important role in bacterial biofilm formation and their production is strongly correlated with pathogenicity and virulence. In many bacteria the biosynthetic machineries required for assembly of these exopolysaccharides are regulated by common signaling molecules, with the second messenger cyclic di-guanosine monophosphate (c - di-GMP) playing an especially important role in the post-translational activation of exopolysaccharide biosynthesis. Research on treatments of antibiotic-resistant and biofilm-forming bacteria through direct targeting of c-di-GMP signaling has shown promise, including peptide-based treatments that sequestermore »intracellular c-di-GMP. In this review, we will examine the direct role c-di-GMP plays in the biosynthesis and export of biofilm exopolysaccharides with a focus on the mechanism of post-translational activation of these pathways, as well as describe novel approaches to inhibit biofilm formation through direct targeting of c-di-GMP.« less
  3. Abstract Background Acinetobacter baumannii is a gram-negative bacterium which causes opportunistic infections in immunocompromised hosts. Genome plasticity has given rise to a wide range of strain variation with respect to antimicrobial resistance profiles and expression of virulence factors which lead to altered phenotypes associated with pathogenesis. The purpose of this study was to analyze clinical strains of A. baumannii for phenotypic variation that might correlate with virulence phenotypes, antimicrobial resistance patterns, or strain isolation source. We hypothesized that individual strain virulence phenotypes might be associated with anatomical site of isolation or alterations in susceptibility to antimicrobial interventions. Methodology A cohort of 17 clinical isolates of A. baumannii isolated from diverse anatomical sites were evaluated to ascertain phenotypic patterns including biofilm formation, hemolysis, motility, and antimicrobial resistance. Antibiotic susceptibility/resistance to ampicillin-sulbactam, amikacin, ceftriaxone, ceftazidime, cefotaxime, ciprofloxacin, cefepime, gentamicin, levofloxacin, meropenem, piperacillin, trimethoprim-sulfamethoxazole, ticarcillin- K clavulanate, tetracyclin, and tobramycin was determined. Results Antibiotic resistance was prevalent in many strains including resistance to ampicillin-sulbactam, amikacin, ceftriaxone, ceftazidime, cefotaxime, ciprofloxacin, cefepime, gentamicin, levofloxacin, meropenem, piperacillin, trimethoprim-sulfamethoxazole, ticarcillin- K clavulanate, tetracyclin, and tobramycin. All strains tested induced hemolysis on agar plate detection assays. Wound-isolated strains of A. baumannii exhibited higher motility than strains isolated frommore »blood, urine or Foley catheter, or sputum/bronchial wash. A. baumannii strains isolated from patient blood samples formed significantly more biofilm than isolates from wounds, sputum or bronchial wash samples. An inverse relationship between motility and biofilm formation was observed in the cohort of 17 clinical isolates of A. baumannii tested in this study. Motility was also inversely correlated with induction of hemolysis. An inverse correlation was observed between hemolysis and resistance to ticarcillin-k clavulanate, meropenem, and piperacillin. An inverse correlation was also observed between motility and resistance to ampicillin-sulbactam, ceftriaxone, ceftoxamine, ceftazidime, ciprofloxacin, or levofloxacin. Conclusions Strain dependent variations in biofilm and motility are associated with anatomical site of isolation. Biofilm and hemolysis production both have an inverse association with motility in the cohort of strains utilized in this study, and motility and hemolysis were inversely correlated with resistance to numerous antibiotics.« less
  4. This study aimed to address the significant problems of bacterial biofilms found in medical fields and many industries. It explores the potential of classic photoactive carbon dots (CDots), with 2,2′-(ethylenedioxy)bis (ethylamine) (EDA) for dot surface functionalization (thus, EDA-CDots) for their inhibitory effect on B. subtilis biofilm formation and the inactivation of B. subtilis cells within established biofilm. The EDA-CDots were synthesized by chemical functionalization of selected small carbon nanoparticles with EDA molecules in amidation reactions. The inhibitory efficacy of CDots with visible light against biofilm formation was dependent significantly on the time point when CDots were added; the earlier the CDots were added, the better the inhibitory effect on the biofilm formation. The evaluation of antibacterial action of light-activated EDA-CDots against planktonic B. subtilis cells versus the cells in biofilm indicate that CDots are highly effective for inactivating planktonic cells but barely inactivate cells in established biofilms. However, when coupling with chelating agents (e.g., EDTA) to target the biofilm architecture by breaking or weakening the EPS protection, much enhanced photoinactivation of biofilm-associated cells by CDots was achieved. The study demonstrates the potential of CDots to prevent the initiation of biofilm formation and to inhibit biofilm growth at an early stage.more »Strategic combination treatment could enhance the effectiveness of photoinactivation by CDots to biofilm-associated cells.« less
  5. Bacterial biofilms represent a basic form of multicellular organization that confers survival advantages to constituent cells. The sequential stages of cell ordering during biofilm development have been studied in the pathogen and model biofilm-formerVibrio cholerae. It is unknown how spatial trajectories of individual cells and the collective motions of many cells drive biofilm expansion. We developed dual-view light-sheet microscopy to investigate the dynamics of biofilm development from a founder cell to a mature three-dimensional community. Tracking of individual cells revealed two distinct fates: one set of biofilm cells expanded ballistically outward, while the other became trapped at the substrate. A collective fountain-like flow transported cells to the biofilm front, bypassing members trapped at the substrate and facilitating lateral biofilm expansion. This collective flow pattern was quantitatively captured by a continuum model of biofilm growth against substrate friction. Coordinated cell movement required the matrix protein RbmA, without which cells expanded erratically. Thus, tracking cell lineages and trajectories in space and time revealed how multicellular structures form from a single founder cell.