Biology:MecA (gene)

The mecA gene is a gene found in bacterial cells which allows a bacterium to be resistant to antibiotics such as methicillin, penicillin and other penicillin-like antibiotics.

The most commonly known carrier of the mecA gene is the bacterium known as Methicillin-resistant Staphylococcus aureus (MRSA). In Staphylococcus species, mecA is spread on the SCCmec genetic element. Resistant strains are responsible for many infections originating in hospitals.

The mecA gene does not allow the ringlike structure of penicillin-like antibiotics to bind to the enzymes that help form the cell wall of the bacterium (transpeptidases), and hence the bacteria is able to replicate as normal. mecA is located on the staphylococcal chromosome cassette mec. The gene encodes the protein PBP2A (penicillin binding protein 2A). PBP2A has a low affinity for beta-lactam antibiotics such as methicillin and penicillin. This enables transpeptidase activity in the presence of beta-lactams, preventing them from inhibiting cell wall synthesis.

History

Methicillin resistance was first exhibited in hospitals where the bacteria Staphylococcus aureus was more aggressive and not responding to the methicillin treatment provided by doctors. Throughout the years, the rate of this strain of S. aureus has continued to increase, reaching up to 60% of British hospitals, and has spread throughout the world and is not restricted to hospital settings. The source of this resistance was determined to be the mecA gene, that was acquired through a mobile genetic element, staphylococcal cassette chromosome mec, which is present in all known methicillin resistant strains of S. aureus. On February 27, 2017, the World Health Organization released a list of priority bacterial resistant pathogens, naming methicillin-resistant S. aureus, as a high priority target for further research and treatment development.

Detection

Successful treatment of methicillin resistant S. aureus begins with the detection and confirmation that the strain in question actually possesses the mecA gene, responsible for the resistance. The use of polymerase chain reaction (PCR) is typically used to detect the presence of the mecA gene, alternative methods can be used as that can be as specific as PCR. Enzymatic detection PCR, which labels the PCR with enzymes, detectable by immunoabsorbant assays, takes less time, and does not require the need for gel electrophoresis, which can be costly, tedious, and unpredictable. The use of cefoxitin disc diffusion test uses phenotypic resistance to test not only for methicillin resistant strains, but also for low resistant strains also The presence of the mecA gene cannot be used on its own to determine resistant strains; further phenotypic assays of mecA positive strains, can determine the level at which the strain is resistant to methicillin. These phenotypic assays cannot rely on the accumulation of Penicillin-binding protein 2a, the protein product of the mecA gene, as a test for methicillin resistance, because there is no connection between amount of protein and resistance for this gene; the determination of level of resistance is still unknown.

Structure

The mecA gene is contained on a mobile gene element, called the staphylococcal cassette chromosome mec, from which the gene can undergo horizontal gene transfer and insert itself into the host species, which can be any species in the genus Staphylococcus. This DNA cassette is a 52 kilobase piece of DNA, that contains the mecA gene, and two recombinase genes, ccrA and ccrB, which the plasmid uses to insert itself into the genome of the host. These recombinases are essential for the proper insertion of the mecA complex into the host genome. Multiple variants of these genes have been isolated from resistant strains of S. aureus, but all of the variants have similar function and the same insertion site, near the host DNA origin of replication. The mecA gene also forms a complex with two regulatory units, mecI and mecR1. These two genes have the capability to repress mecA, deletions or knock-outs in these genes show an increase in resistance of S. aureus to methicillin. The S. aureus strains isolated from humans either lack these regulatory elements, or contain mutations in these genes that cause a loss of function of the protein products that inhibit mecA. This in turn, causes mecA to be constitutive transcription of the mecA. This cassette chromosome shows the ability to move across species. Two other Staphylococci species, S.epidermidis and S.haemolyticus, show conservation in this insertion site, but it is not limited to the mecA gene, but also other non-essential genes that can be carried by the cassette chromosome.

Mechanism of resistance

Penicillin, its derivatives and methicilin, are beta-lactams that all work by disrupting the mechanical structure of the bacterial cell wall by reacting with the cell wall forming penicillin-binding protein family (PBP 1,2, 3 and 4), causing the cytoplasm to leak and the cell to die. However, the mecA gene codes for PBP2a, that has a lower affinity for antibiotic drugs, which keeps the structural integrity of the cell wall, preventing cell death. The synthesis of the bacterial cell wall in S. aureus is dependent on transglycosylation, to form linear polymer of sugar monomers, and transpeptidation, to form an interlinking peptides to strengthen the newly developed cell wall. PBPs have a transpeptidase domain, but transglycosylation was thought to only be carried out by monofunctional enzymes, however PBP2 has domains to carry out both essential processes. When antibiotics are introduced to the medium, they bind to the transpeptidation domain and inhibit the ability of PBPs to cross-link muropeptides, therefore preventing the formation of stable cell wall. However, with cooperative action, PBP2a lacks the proper receptor for the antibiotics and continues the transpeptidation, preventing the breakdown of the cell wall. The functionality of PBP2a is dependent on two structural factors on the cell wall of S. aureus. First, there in order for PBP2a to properly fit onto the cell wall, to continue transpeptidation, requires proper amino acid residues, specifically a pentaglycine residue and an amidated glutamate residue. Furthermore, PBP2a has an effective transpeptidase activity, but lacks a transglycosylation domain of PBP2, which builds the backbone of the cell wall with polysaccharide monomers, and so PBP2a must rely on PBP2 to continue this process. The latter of these factors is an enzymatic reaction that can be targeted to improve the ability of beta-lactams to prevent cell wall synthesis in resistant S. aureus. Identification of genetic inhibitors of glycosylases involved in the cell wall synthesis, and modulating the expression of these inhibitors can resensitize these previously resistant bacteria to beta-lactam treatment. for example, Epicatechin gallate, a compound found in green tea, has shown signs of lowering the resistance to beta-lactams, to the point where oxacillin becomes effective to inhibit the formation of the cell wall, by inhibiting PBP2 and PBP2a.

Evidence has also show that interactions with other genes have been identified to decrease resistance to beta-lactams in resistant strains of S. aureus. The gene networks have been identified, are mainly involved in cell division, and cell wall synthesis and function, where there PBP2a localizes. Furthermore, PBP2a is not the only protein from the PBP family to affect the resistance of S. aureus to antibiotics. PBP4 showed a to be helpful in maintaining oxacillin resistance, as oxacillin resistance was lowered in S. aureus strains when expression of both PBP4 was inhibited but PBP2a was not.

Evolutionary history

The mecA gene is acquired and transmitted through a mobile genetic element, that inserts itself into the host genome. Evidence shows that there is a conservation of structure between the mecA gene product and a homologous mecA gene product in the bacteria Staphylococcus sciuri. Currently, there is no known function for the mecA homologue in S. sciuri but the evidence points towards these genes as being a precursor for the mecA gene found in S. aureus. Furthermore, the structure of the protein product of this homologue is so similar that it can be used in S. aureus. When the mecA homologue of beta lactam resistant S. sciuri is inserted into antibiotic sensitive S. aureus, there is an increase in the resistance to antibiotics. Even though there is a difference in the muropeptides that both species use, the protein product of the S. sciuri mecA gene is still able to continue cell wall synthesis when the PBP protein family is inhibited by a beta lactam.

In order to further understand the origin of the mecA gene, specifically the mecA complex found on the Staphylococcal cassette chromosome, researchers used the mecA gene from S. sciuri in comparison to other Staphylococci species. After nucleotide analysis, the sequence of the mecA gene is almost identical to the mecA homologue found in Staphylococcus fleurettii, the most significant candidate for the origin of the mecA gene on the staphylococcal cassette chromosome. Since this gene is contained in the genome of S. fleurettii, there is no way to justify this species to create the cassette chromosome.