**3.2 Slow cell growth in biofilms**

Slow cell growth of the bacterial has been found in mature biofilms [17]. This phenomenon is responsible for the decreased susceptibility of bacteria in biofilms to antibiotics requiring growing organisms for their bactericidal effects. For example, penicillins and cephalosporins prefer to killing the growing bacterial cells, and the rate of killing cells is proportional to the growth rate [17]. It is well known that most antimicrobial agents act on certain types of macromolecular synthesis to exert antimicrobial activities, such as the synthesis of enzymes, proteins, and nucleic acids (DNA or RNA). Thus, these antibiotics have little effects on bacteria with stagnant macromolecular synthesis, which leads to bacterial drug resistance.

Nutrition restriction is one of reasons that are responsible for slow cell growth. The mechanism of nutrition restriction is closely related to the osmotic restriction. Due to the existence of biofilm osmotic restriction, nutrients are not easy to pass through biofilm, which leads to the lack of nutrition in biofilm and slows down the growth rate of inner layer bacteria. This slow growth state of inner layer bacteria also forms a protective mechanism, which reduces the susceptibility of bacteria to antibiotics [24].

When the biofilm cells are exposed to antibiotics, the bacteria on the surface of the biofilm are killed by the drug, and the cells in the middle and deep layers of the biofilm are not affected. After the antibiotic treatment stops, the remaining bacteria will use dead bacteria as nutrients to reproduce rapidly, which can only take a few hours to reproduce [25, 26].

**105**

*Formation, Antibiotic Resistance, and Control Strategies of* Staphylococcus epidermidis *Biofilm*

Delayed penetration of the antibiotics through the biofilm matrix and slow rate of bacterial reproduction in biofilm cannot explain entirely the resistance of biofilms to one important class of antibiotics, namely the fluoroquinolones. This class of antimicrobial agents equilibrates across bacterial biofilms and exerts bactericidal effect on nondividing cells [17]. Although a dose-dependent bactericidal action was observed in *P. aeruginosa* biofilms by the fluoroquinolones ofloxacin and ciprofloxacin, a further increase in the antibiotic concentration or a prolonged drug action period did not improve killing rates after an initial 3- to 4-log drop bacterial counts. This result suggested that a small portion of "persister" cells occurs after administration of fluoroquinolones [17, 27, 28]. The most significant difference between persisters and mutant resistant strains is that the drug resistance of persisters is only a phenotypic variation without gene mutation, so this phenotype is not genetic. These strains were collected, recultured, and detected the drug resistance. It was intriguing that the drug resistance disappeared, and the minimum inhibitory concentrations (MICs) were the same level as those of parent strains. Meanwhile, the resistant strains caused by mutation showed a stable genetic drug resistance,

Persister cells in biofilms are considered to the key in the extraordinary survival properties of biofilms. The dynamic features of biofilm formation and shedding of cells from one biofilm to form a new biofilm may also explain the chronic nature of biofilm infections and the need for extending antimicrobial agent treatment to

Because the expression of toxins and other virulence factors is less in *S. epidermidis*, the biofilm forming capacity is its major virulence factor. Biofilm growth is characterized by high resistance to antimicrobial agents and host immune responses, making biofilm eradication tremendously difficult. The increasing prevalence of multidrug-resistant *S. epidermidis* strains additionally hampers antimicrobial therapy. Therefore, targeting factors expressed at different phases in biofilm formation might offer new tools to combat *S. epidermidis* infections.

The first step of biofilm formation is bacterial adherence to the host cell surface. Direct binding to host cell surface is mediated by electrostatic and hydrophobic interactions and van der Waals forces and affected by physicochemical variables [29]. Found in our research, after investigating the antibiofilm activities of spent media from 185 Actinomycete strains using two *S. epidermidis* strains (ATCC 35984 and a clinical strain 5-121-2) as target bacteria, three strains of tested Actinomycete (TRM 46200, TRM 41337, and TRM 46814) showed a significant inhibition against *S. epidermidis* biofilm formation without affecting the growth of planktonic cells. Effect of Actinomycete supernatants on cell surface hydrophobicity (CSH) of *S. epidermidis* was measured by Microbial Adhesion to Hydrocarbon (MATH) assay. The adhesion of staphylococci to n-hexadecane was used to measure the hydrophobicity of *S. epidermidis*. All the crude proteins from spent media showed a reduction in the CSH against *S. epidermidis* ATCC 35984 and 5-121-2, which explain at least in part the inhibitory effect of Actinomycete supernatants on biofilm reduction [19].

*DOI: http://dx.doi.org/10.5772/intechopen.89800*

and MICs were higher than those of parent strains [28].

**4. Control strategies to** *S. epidermidis* **biofilm formation**

disturb the dynamics of biofilm formation [17].

**4.1 Inhibition of initial attachment**

**3.3 Formation of persister cells**

*Formation, Antibiotic Resistance, and Control Strategies of* Staphylococcus epidermidis *Biofilm DOI: http://dx.doi.org/10.5772/intechopen.89800*

### **3.3 Formation of persister cells**

*Bacterial Biofilms*

responsible for drug resistance.

tance of *S. epidermidis* biofilms.

**3.2 Slow cell growth in biofilms**

**3.1 Antibiotic penetration of biofilms**

*S. epidermidis* and other bacterial species produce an extracellular matrix called glycocalyx or slime, which is a highly hydrated complex composed of teichoic acids, proteins, and exopolysaccharides. In biofilms, poor antibiotic penetration, nutrient limitation and slow growth, and formation of persister cells are hypothesized to be

Biofilms are typically characterized by dense, highly hydrated clusters of bacterial cells enclosed in a self-produced polymeric matrix that is primarily composed of exopolysaccharides such as polysaccharide intercellular adhesin (PIA) in staphylococci and adherent to a surface. This matrix, also termed slime or extracellular polymeric substance (EPS), impairs the access of antimicrobial agents to the bacterial cells [21]. Additionally, either a reaction of EPS with or its adsorption to the components of the biofilm matrix can delay penetration of the antibiotics through the biofilm matrix. The effective diffusion coefficients of solutes in biofilms average about 40% of the respective diffusion coefficient in pure water [20]. *S. epidermidis* slime has been found to remarkably decrease the activity of the glycopeptides vancomycin and teicoplanin. The efficacy of cloxacillin, amoxicillin/clavulanic acid, imipenem, cefpirome, erythromycin, roxithromycin, clindamycin, fusidic acid, trimethoprim/sulfamethoxazole, doxycycline, gentamicin, tobramycin, netilmicin, amikacin, isepamicin, ofloxacin, ciprofloxacin, and daptomycin is also moderately affected by the exopolysaccharide matrix of *S. epidermidis*. Other studies have suggested that *S. epidermidis* glycocalyx reduces susceptibility to pefloxacin and moderately affects the activity of daptomycin, linezolid, and quinupristin/dalfopristin [22, 23]. The role of biofilm matrix in retarding the penetration of antibiotics is thereby contributed to the drug resis-

Slow cell growth of the bacterial has been found in mature biofilms [17]. This phenomenon is responsible for the decreased susceptibility of bacteria in biofilms to antibiotics requiring growing organisms for their bactericidal effects. For example, penicillins and cephalosporins prefer to killing the growing bacterial cells, and the rate of killing cells is proportional to the growth rate [17]. It is well known that most antimicrobial agents act on certain types of macromolecular synthesis to exert antimicrobial activities, such as the synthesis of enzymes, proteins, and nucleic acids (DNA or RNA). Thus, these antibiotics have little effects on bacteria with stagnant

Nutrition restriction is one of reasons that are responsible for slow cell growth. The mechanism of nutrition restriction is closely related to the osmotic restriction. Due to the existence of biofilm osmotic restriction, nutrients are not easy to pass through biofilm, which leads to the lack of nutrition in biofilm and slows down the growth rate of inner layer bacteria. This slow growth state of inner layer bacteria also forms a protective mechanism, which reduces the susceptibility of bacteria to

When the biofilm cells are exposed to antibiotics, the bacteria on the surface of the biofilm are killed by the drug, and the cells in the middle and deep layers of the biofilm are not affected. After the antibiotic treatment stops, the remaining bacteria will use dead bacteria as nutrients to reproduce rapidly, which can only take a few

macromolecular synthesis, which leads to bacterial drug resistance.

**104**

antibiotics [24].

hours to reproduce [25, 26].

Delayed penetration of the antibiotics through the biofilm matrix and slow rate of bacterial reproduction in biofilm cannot explain entirely the resistance of biofilms to one important class of antibiotics, namely the fluoroquinolones. This class of antimicrobial agents equilibrates across bacterial biofilms and exerts bactericidal effect on nondividing cells [17]. Although a dose-dependent bactericidal action was observed in *P. aeruginosa* biofilms by the fluoroquinolones ofloxacin and ciprofloxacin, a further increase in the antibiotic concentration or a prolonged drug action period did not improve killing rates after an initial 3- to 4-log drop bacterial counts. This result suggested that a small portion of "persister" cells occurs after administration of fluoroquinolones [17, 27, 28]. The most significant difference between persisters and mutant resistant strains is that the drug resistance of persisters is only a phenotypic variation without gene mutation, so this phenotype is not genetic. These strains were collected, recultured, and detected the drug resistance. It was intriguing that the drug resistance disappeared, and the minimum inhibitory concentrations (MICs) were the same level as those of parent strains. Meanwhile, the resistant strains caused by mutation showed a stable genetic drug resistance, and MICs were higher than those of parent strains [28].

Persister cells in biofilms are considered to the key in the extraordinary survival properties of biofilms. The dynamic features of biofilm formation and shedding of cells from one biofilm to form a new biofilm may also explain the chronic nature of biofilm infections and the need for extending antimicrobial agent treatment to disturb the dynamics of biofilm formation [17].
