1) (Defining) antibiotic resistanceAny new therapeutic agent is at risk of developing some form of tolerance or resistance from the time it is first used 1. The most prominent example of this is antibiotic resistance which is the process by which a bacteria develops resistance to an antimicrobial drug, which it was previously susceptible to. Different classes of antibiotics exist and have specific modes of action by which they inhibit the growth of bacteria or kill the microorganisms. These include inhibition of cell wall synthesis, protein synthesis, DNA synthesis, RNA synthesis, inhibition of a metabolic pathway and cell membrane disorganisation 2. Bacteria are regularly remodelling their cell walls, synthesising and breaking down peptidoglycan as they grow and divide. Penicillin-binding proteins are a sub-group of the DD-transpeptidases and are involved in the final stages of peptidoglycan synthesis, these enzymes catalyse the polymerisation of the glycan strand and the cross-linking between glycan chains. ?-lactam antibiotics, such as penicillin, contain a four-membered ?-lactam ring, which is responsible for inhibiting the synthesis of peptidoglycan. The ?-lactam region of the drug mimics the D-Ala-D-Ala end of the peptide to which the DD-transpeptidase enzyme binds to. Penicillin is able to bind irreversibly to the active site of the enzyme, preventing it from cross-linking the peptidoglycan strands. As a consequence, there is an imbalance between cell wall production and degradation and the cell wall loses its integrity, causing cell lysis 3.?All antibiotics work by binding to specific bacterial molecular targets such as enzymes or organelles. Bacteria can thus become resistant to these drugs by developing mechanisms to prevent them from binding to their molecular target. Bacteria have developed resistance to all classes of antibiotics on the market. Some bacteria have even developed simultaneous resistance to different types of antibiotics, creating dangerous multidrug-resistant (MDR) bacterial strains. Multidrug resistance has been discovered in many clinically important species such as Pseudomonas aeruginosa, Acinetobacter baumannii, E. coli, and Klebsiella pneumoniae. A recent database has recorded more than 20,000 potential resistance genes (R genes) of nearly 400 different types, which was estimated from available bacterial genome sequences 1. So, bacteria eventually become resistant to antibiotics but certain anthropogenic factors have contributed to the current increase we see in resistance.2) Factors promoting antibiotic resistance?Multiple factors have contributed to antibiotic resistance since their use in modern medicine. Humans have applied a selection pressure, which has increased the number of resistance genes which may then spread between bacteria. The overuse and overprescription of antibiotics is considered to be the biggest contributing factor to the evolution of resistance. It has been estimated that in 30-50% of cases, the choice, indication and duration of the antibiotic is inaccurate. Even in 1945, Alexander Fleming was concerned with the ramifications of overusing antibiotics and said “the public will demand the drug and … then will begin an era … of abuses” 4. Recent epidemiological studies have established a direct correlation between the consumption of antibiotics and the emergence and spread of resistance genes. Global consumption of antibiotics in human medicine has increased by nearly 40% between 2000 and 2010. There are 3 main strands of evidence that point to resistance emerging as a consequence of antibiotic use. These include laboratory evidence, ecological studies and individual level data. ?It has also been shown that when prescribed antibiotics, many patients do not fulfil the treatment and stop using once they deem their infection is gone, rather than completing their course. In some countries, such as the United Arab Emirates, antibiotics can be bought over the counter. They may not necessarily know what bacterial infection they have acquired or even if they have a bacterial infection at all and yet are still allowed to purchase them. There may be poor infection control in hospitals, poor hygiene and sanitation practices and a lack of rapid laboratory tests which leads to an increase in resistant genes. Antibiotics are also widely used in livestock and fish farming. Strains of resistant bacteria can build up on farms and the resistance is then transferred to bacteria found naturally in our bodies.3) Mechanisms of antibiotic resistanceOnce an antibiotic is used as a therapeutic agent, it is only a matter of time until a resistant pathogen appears. Bacteria have short generation times and large populations and since the average rate of mutation is 1 in 107, then a colony of 1010 bacteria would have mutations in approximately 1000 loci 2. New mutations can increase genetic diversity and this diversity in turn can lead to rapid evolution. The mutations may confer resistance to an antibiotic, allowing the bacteria to persist and reproduce. Resistance to penicillin was first recorded only 2 years after it was introduced into human medicine 2. Pathogens have many different mechanisms of resistance to antibiotics. There are 2 types of resistance which exist; innate intrinsic resistance is where resistance to a particular antibiotic or group of antibiotics is characteristic for a specific bacterial genus, species or entire bacterial group. It may be the result of the lack of a target for the particular antibiotic or because the drug can’t access its target. Acquired resistance is where most isolates of a bacterial species would be fully susceptible to the particular antibiotic, but resistance may arise in a few, or in some cases in many isolates. It may arise through mutation of a chromosomal gene, acquisition of genes by bacteriophages, plasmids, integrons or transposons 5.A common mechanism is the presence of efflux pumps which are transport proteins that pump antibiotics out of the cell into the environment faster than the antibiotic can diffuse in. These are found in both Gram-positive and Gram-negative bacteria. Efflux systems have been discovered in a number of clinically relevant bacteria, such as E. coli (AcrAB-TolC, AcrEF-TolC, EmrB, EmrD9), P. aeruginosa (MexAB-OprM, MexCD-OprJ, MexEF-OprN and MexXY-OprM9), S. pneumoniae (PmrA10) and S. aureus (NorA12) 6.??Pathogens may also inactivate the drug using enzymes that modify or degrade the drug. They alter the drug target site preventing the drug from binding to its target. Gram-negative bacteria produce ?-lactamases which are able to hydrolyse the amide bond of the ?-lactam ring. Gram-positive modify their target site, the penicillin-binding proteins to confer resistance to these antibiotics. Alternatively, bacteria may prevent the drug from entering the cell by modifying membrane permeability or transport systems. Hydrophilic molecules, for example, ?-lactams and tetracyclines, use porins to cross the membrane and are therefore notably affected by changes in membrane permeability. Mutations in the gene encoding porin OprD, normally involved in the transport of antibiotics into the bacterial cell, have been discovered in clinical isolates of Pseudomonas aeruginosa, and confer resistance to carbapenems 7.?Antibiotic resistance genes are present on chromosomes and plasmids and can be transferred between bacteria by vertical or horizontal transmission. Antibiotics can provide a selection pressure and cause the evolution and spread of antibiotic resistance genes.?4) The spread of antibiotic resistanceVertical transmission involves the spontaneous mutation of a cell causing the antibiotic to be ineffective against this strain. The resistant cell will then multiply, increasing the number of resistant microorganisms in the population. Horizontal gene transfer involves the transfer of genetic material between mature cells without having to undergo replication, and it is the main mechanism for the spread of antibiotic resistance genes. There are 3 processes by which this can occur: transformation, transduction and conjugation. ??Natural transformation is the uptake of DNA from medium and is mediated by competence proteins, but this only occurs in approximately 1% of species 8, for example in Staphylococcus, Streptococcus, Pseudomonas, Neiserria. The cell is described as recombinant as there was an exchange of DNA material. In transduction, the resistance gene is integrated into the new host cell chromosome or plasmid along with phage DNA. Bacteriophages move genes from one bacteria to another. In conjugation, the resistance gene moves with the replicating plasmid into a new cell, it is mediated by a pilus that attaches from the donor strain to the recipient strain. ??A previously harmless strain of Streptococcus pneumoniae can cause an infection if the bacteria is placed in a medium containing DNA from a pathogenic strain as it takes up the allele for pathogenicity and exchanges its own allele by the process of natural transformation. Judging from the high number of lysogenic phages present in pneumococcal genomes, transduction may also contribute significantly to the genetic diversity in this species. Several conjugative transposons have been discovered in S. pneumoniae. They often carry tetracycline and erythromycin resistance genes. In fact, it has been found that drug resistance determinants are more frequently found on conjugative transposons than on plasmids 9.5) The global antibiotic resistance crisisAntibiotic resistance is present in every country and is a growing worldwide crisis. Infections which are caused by drug-resistant microorganisms threaten patient safety with worse clinical outcomes and some infections may even be lethal. They also cost health-care systems more than patients infected with non-resistant strains of the same bacteria, one study in the US found that antibiotic resistant pathogens could increase the cost by approximately $6,000 – $30,000 10. This has a huge impact on the NHS, specifically costing millions every year. In the European Union, the CDC found that antibiotic resistance causes 25,000 deaths per year and 2.5 million extra hospital days 11. The cost is approximately €1.5 billion per year due in part to 600 million days of lost productivity. The World Economic Forum in its 2013 Global Risks Report put antimicrobial resistance on a par with the unrestricted proliferation of weapons of mass destruction and global economic meltdown. If nothing is done, it is estimated that antibiotic resistant pathogens will claim 10 million lives by 2050.??In February 2017, the World Health Organisation published a list of 12 families of bacteria for which new antibiotics are needed as they pose a great threat to human health. This list was divided into 3 categories consisting of critical, high and medium priority. The list aims to motivate governments to put in place policies that encourage research and development into the discovery of new antibiotics, by publicly funded agencies and the private sector. The critical priority group contains multidrug resistant bacteria that are particularly prevalent in hospitals, and other healthcare settings, and among patients that require devices such as ventilators and blood catheters. They include Acinetobacter baumannii, Pseudomonas aeurginosa and Enterobacteriaceae species (including Klebsiella, E. coli). These microorganisms may cause severe and sometimes deadly infections, such as pneumonia. These bacteria have developed resistance to most antibiotics found on the market today, including carbapenems and third generation cephalosporins which are used for treating multi-drug resistant bacteria. The high and medium priority bacteria include other increasingly drug-resistant bacteria that cause more common diseases such as gonorrhoea (Neisseria gonorrhoeae)12.