**2. Mechanisms of antibiotic resistant**

#### **2.1 Methicillin resistant** *Staphylococus aureus* **(MRSA)**

Alexander Flemming introduced the antibiotic, Penicillin in 1940s for the treatment of bacterial infection. At that time, *S. aureus* infections were well controlled. However, with the widespread use of this antibiotic in the 1950s, Penicillin-resistant *S. aureus* appeared. It produces pencillinase enzyme, which can hydrolyze the betalactum ring of Penicillin. In 1959, substitution of the natural aminoadipoyl chain from Penicillin with bulkier moieties, developed a semi-synthetic Penicillin, named Methicillin. However, it was not widely used because of its toxicity. It was replaced by similar, more stable Penicillins like Oxacillin, Flucloxacillin and Dicloxacilllin. These antibiotics show good antibacterial activity and are resistant to beta-lactamase substrate. In 1961 the British scientist, Jevons isolated the penicillin stable resistant *S. aureus*. However, the name Methicillin resistant *S. aureus* (MRSA) continues to be used [9].

Bifunctional Transglycolylase-transpeptidase (Penicillin binding protein 'a' or PBPa) is the inhibitory target of beta-lactam antibiotics in *S. aureus*. The transglycolylase domain is responsible for transferring the disaccharide pentapeptide (L-alanine, D-glutamine, Lysine and 2 D-alanines) from membrane bound lipid to growing chains of polysaccharide. Domain of Transpeptidase (TP) cross-links the glycine bridge and links the D-alanine of 4th position to adjacent chain of peptidoglycan layer to make the cell-wall strong. The active site of Transpeptidase (TP) serine is blocked (i.e., PBP2a) by causing structural analogous changes of D-Ala4 to D-Ala5. This leads to breakdown of beta lactam ring and a penicilloyl-O-serine intermediate is formed [10].

PBP2a is encoded by mec A gene. This mecA gene is a mobile genetic element integrated into the chromosomal element (SCCmec) of Methicillin sensitive *S. aureus*. The mecA gene is transfered to other *S. aureus* via horizontal gene transfer mechanisms. Resistance confered by mec A gene is broad spectrum and shows resistance to all beta-lactum antibiotics except Ceftaroline and Ceftobiprole [11].

SCCmec contains two essential components such as mec gene complex and ccr gene complex. The mec gene complex contains mecA and is associated with regulatory and insertion sequences. It has been classified into 6 different classes (A, B, C1, C2, D and E) along with ccr complex (Cassette chromosome recombinase) genes. It encodes for the enzyme, 'recombinase' that helps in integration and excision of SCCmec into the chromosome. There are 3 different types of recombinase enzymes, namely ccrA, ccrB, and ccrC. Recombinase enzymes are further classified into eight different types based on the existing recombinase and allotypes in the different characteristics.

SCCmecs are classified into 8 types and subtypes according to 'International Working Group on the Staphylococcal Cassette Chromosome elements' [12].
