**3. Mechanisms of resistance to antimicrobial agents**

The ability of an organism to overcome the action of an antimicrobial agent to which it was previously susceptible is a general definition of antimicrobial resistance [39]. With the growing production of MDR strains (i.e. resistance to at least three antibiotics), nosocomial infection caused by antibiotic resistant *P. aeruginosa* has emerged as a major concern in clinical care settings [40]. Because of its outer membrane with low permeability (1/100 of the permeability of the outer membrane of *E. coli*), *P. aeruginosa* exhibits intrinsic resistance to various antimicrobial agents (β-lactam and penem group of antibiotics) [41, 42]. While several other processes are also responsible for their intrinsic resistance, including the efflux system that expels antibiotics from the cell's bacteria and the production of inactivating enzyme antibiotics. This bacterium, however is a highly diverse pathogen capable of adapting to the conditions around it. When subjected to selective pressure from antibiotics, the mediated reaction encourages bacterial survival and improves resistance to antibiotics [43–45].

The development of antibiotic resistance during host colonization of patients with CF has been confirmed, with *P. aeruginosa* strains developing and gaining resistance during antimicrobial therapy [46]. Studies have shown a clear link between increased applications of ciprofloxacin, with a growing incidence of strains resistant to ciprofloxacin [47]. Therefore the excessive use of antimicrobial agents is another factor associated with the rise in MDR-*Psedomonas aeruginosa.* This acquired resistance may be attributable to the effects of the mutational event or the acquisition by horizontal gene transfer of the resistance gene and may occur during the mutational event of antibiotic therapy, leading to over-expression of endogenous β-lactamases or efflux pump, specific porin expression [48].

#### **3.1 Resistance to β-lactam**

Inhibition of the synthesis of the bacterial peptidoglycan cell wall requires β-lactam antibiotics [39]. Penicillin, cephalosporin, carbapenem and monobactam are included in this class. These classes include piperacillin and ticarcillin (penicillin), ceftazidime (cephalosporin 3rd generation), cefepime (cephalosporin 4th generation), aztreonam (monobactam), imipenem, meropenem and doripenem (carbapenems) are most powerful β-lactam widely used to treat *P. aeruginosa* is β-lactam [49]. These enzymes break the amide bond of the β-lactam ring through

**81**

available β-lactam.

**3.4 Resistance to aminoglycoside**

**3.3 Class A carbenicillin hydrolysing β-lactamases**

*Chemotherapy and Mechanisms of Action of Antimicrobial Agent*

resistance to the β-lactam mediated by the action of β-lactamases, rendering the antimicrobial ineffective. The expression of endogenous β-lactamases or the expression of acquired β-lactamases may be due to this inactivation of the drug. To date, hundreds of β-lactamases have been recognized and are distinguished by their substrate specificity. There are four major groups of beta-lactamases known in *P. aeruginosa* on the basis of Amber's molecular classification system: A-D [50]. Through the serine-residue catalytic activity, classes A, C and D inactivate the β-lactams, while

class B or metallo- β-lactamases (MBLs) require zinc for their action [51].

coded AmpC β-lactamase (cephalosporinase) belonging to the Ambler-based molecular class C and the first functional group according to Bush et al. [54, 55]. However, production of chromosomal cephalosporin in *P. aeruginosa*, in the presence of inducing β-lactams (especially imipenem), can increase from 100 to 1000 times [56]. β-lactamase inhibitors used in clinical practice, such as clavulanic acid, sulbactam and tazobactam, do not inhibit AmpC cephalosporinase function. β-lactamase of AmpC is encoded by the gene ampC [57, 58]. Several genes, including ampR, ampG, and ampD, are involved in ampC gene induction. AmpR encodes a positive transcriptional regulator and this regulator is required for the induction of β-lactamase. AmpG, a transmembrane protein that functions as a permease for 1,6-anhydromurapeptides, which are known to be the signal molecules involved in the induction of ampC, is the second gene involved. The third gene, ampD, encodes a cytosolic amidase of N-acetyl-anhydromuramyl-L-alanine that hydrolyses 1,6-anhydromurapeptides, which functions as an ampC expression repressor. The 4th chromosome, ampE, encodes the protein of the cytoplasmic membrane that serves as the molecule of the sensory transducer necessary for induction. Except for avibactam, the activity of this AmpC β-lactamase is not inhibited by commercially

Four β-lactamases (PSE- of *Pseudomonas* specific enzyme) carbenicillin hydrolyzing enzymes were identified in *P. aeruginosa*: PSE-1 (CARB-2), PSE-4 (CARB-1), CARB-3 and CARB-4 [59]. These enzymes belong to the group of β-lactamases of molecular class A and include carboxypenicillins, ureidopenicillins and cefsulodine in their substrate profile. These enzymes belong to functional group 2c and molecular class A [60]. Commercially available β-lactam inhibitors, such as clavulanic acid,

Aminoglycosides are a microbial protein synthesis inhibitor which act by binding to the ribosomal subunit of the bacterial 30S and interfering with the initiation of protein synthesis. Resistance to aminoglycosides in *Pseudomonas* is mediated by

tazobactam, and sulbactam, can inhibit the activity of this β-lactamase [61].

In particular, the development of endogenous β-lactamase, such as chromosomal cephalosporinase (AmpC β-lactamase). A variety of β-lactams, such as benzyl penicillin, narrow spectrum cephalosporin and imipenem, can be induced in *P. aeruginosa*. Naturally, *P. aeruginosa* is susceptible to carboxypenicillins, ceftazidime and aztreonam, but it can develop resistance through a mutation in the gene that contributes to AmpC β-lactamase hyper-production [52, 53]. The enzyme is usually produced in small amounts ('low-level' expression), resistance to aminopenicillins and to most early cephalosporins is determined. *P. aeruginosa* produces an inducible chromosome-

*DOI: http://dx.doi.org/10.5772/intechopen.95476*

**3.2 AmpC β-lactamase (Cephelosporinase)**

*Chemotherapy and Mechanisms of Action of Antimicrobial Agent DOI: http://dx.doi.org/10.5772/intechopen.95476*

*Pseudomonas aeruginosa* - Biofilm Formation, Infections and Treatments

it inhibits tetrahydrofolate synthesis when bound to this enzyme [29].

**3. Mechanisms of resistance to antimicrobial agents**

By competitively blocking the biosynthesis of tetrahydrofolate, which acts as a carrier of one-carbon fragments and is required for the ultimate synthesis of DNA, RNA and bacterial cell wall proteins, trimethoprim and sulfonamides interfere with folic acid metabolism in the microbial cell. Bacteria and protozoan parasites typically lack a transport mechanism in order to extract preformed folic acid from their host, unlike mammals [29]. Most of these species, while some are capable of using exogenous thymidine, must synthesize folic acid, circumventing the need for metabolism of folic acid. The conversion of pteridine and p-aminobenzoic acid (PABA) to dihydrofolic acid by the pteridine synthetase enzyme is competitively inhibited by sulfonamides. Sulfonamides have a greater affinity for pteridine synthetase than for PABA. Trimethoprim has a huge affinity (10,000 to 100,000 times greater than that of the mammalian enzyme) for bacterial dihydrofolate reductase;

The ability of an organism to overcome the action of an antimicrobial agent to which it was previously susceptible is a general definition of antimicrobial resistance [39]. With the growing production of MDR strains (i.e. resistance to at least three antibiotics), nosocomial infection caused by antibiotic resistant *P. aeruginosa* has emerged as a major concern in clinical care settings [40]. Because of its outer membrane with low permeability (1/100 of the permeability of the outer membrane of *E. coli*), *P. aeruginosa* exhibits intrinsic resistance to various antimicrobial agents (β-lactam and penem group of antibiotics) [41, 42]. While several other processes are also responsible for their intrinsic resistance, including the efflux system that expels antibiotics from the cell's bacteria and the production of inactivating enzyme antibiotics. This bacterium, however is a highly diverse pathogen capable of adapting to the conditions around it. When subjected to selective pressure from antibiotics, the mediated reaction encourages bacterial survival and improves resistance to

The development of antibiotic resistance during host colonization of patients with CF has been confirmed, with *P. aeruginosa* strains developing and gaining resistance during antimicrobial therapy [46]. Studies have shown a clear link between increased applications of ciprofloxacin, with a growing incidence of strains resistant to ciprofloxacin [47]. Therefore the excessive use of antimicrobial agents is another factor associated with the rise in MDR-*Psedomonas aeruginosa.* This acquired resistance may be attributable to the effects of the mutational event or the acquisition by horizontal gene transfer of the resistance gene and may occur during the mutational event of antibiotic therapy, leading to over-expression of endogenous β-lactamases or efflux pump, specific porin expression [48].

Inhibition of the synthesis of the bacterial peptidoglycan cell wall requires β-lactam antibiotics [39]. Penicillin, cephalosporin, carbapenem and monobactam are included in this class. These classes include piperacillin and ticarcillin (penicillin), ceftazidime (cephalosporin 3rd generation), cefepime (cephalosporin 4th generation), aztreonam (monobactam), imipenem, meropenem and doripenem (carbapenems) are most powerful β-lactam widely used to treat *P. aeruginosa* is β-lactam [49]. These enzymes break the amide bond of the β-lactam ring through

**2.6 Inhibition of microbial metabolic pathways**

**80**

antibiotics [43–45].

**3.1 Resistance to β-lactam**

resistance to the β-lactam mediated by the action of β-lactamases, rendering the antimicrobial ineffective. The expression of endogenous β-lactamases or the expression of acquired β-lactamases may be due to this inactivation of the drug. To date, hundreds of β-lactamases have been recognized and are distinguished by their substrate specificity. There are four major groups of beta-lactamases known in *P. aeruginosa* on the basis of Amber's molecular classification system: A-D [50]. Through the serine-residue catalytic activity, classes A, C and D inactivate the β-lactams, while class B or metallo- β-lactamases (MBLs) require zinc for their action [51].
