Molecular Mechanism of Drug resistance
Drug resistance is the reduction in effectiveness of a drug in curing a disease or improving a patient’s symptoms. When the drug is not intended to kill or inhibit a pathogen, then the term is equivalent to dosage failure or drug tolerance. More commonly, the term is used in the context of diseases caused by pathogens.Pathogens are said to be drug-resistant when drugs meant to neutralize them have reduced effect. When an organism is resistant to more than one drug, it is said to be multidrug resistant.Drug resistance is an example of evolution in microorganisms. Individuals that are not susceptible to the drug effects are capable of surviving drug treatment, and therefore have greater fitness than susceptible individuals. By the process of natural selection, drug resistant traits are selected for in subsequent offspring, resulting in a population that is drug resistant.Multiple drug resistance or Multidrug resistance is a condition enabling a disease-causing organism to resist distinct drugs or chemicals of a wide variety of structure and function targeted at eradicating the organism. Organisms that display multidrug resistance can be pathologic cells, including bacterial and neoplastic (tumor) cells.Cross-resistance is the tolerance to a usually toxic substance as a result of exposure to a similarly acting substance. It is a phenomenon affecting e.g. pesticides and antibiotics.as an example rifabutin and rifapin cross react in the treatment of tuberculosis. Various microorganisms have survived for thousands of years by their being able to adapt to antimicrobial agents. They do so via spontaneous mutation or by DNA transfer. It is this very process that enables some bacteria to oppose the assault of certain antibiotics, rendering the antibiotics ineffective. These microorganisms employ several mechanisms in attaining multidrug resistance:
- No longer relying on a glycoprotein cell wall
- Enzymatic deactivation of antibiotics
- Decreased cell wall permeability to antibiotics
- Altered target sites of antibiotic
- Efflux mechanisms to remove antibiotics
- Increased mutation rate as a stress response
Many different bacteria now exhibit multidrug resistance, including staphylococci, enterococci, gonococci, streptococci, salmonella, Mycobacterium tuberculosis and others. In addition, some resistant bacteria are able to transfer copies of DNA that codes for a mechanism of resistance to other bacteria, thereby conferring resistance to their neighbors, which then are also able to pass on the resistant gene.
To limit the development of antibiotic resistance, one should:
- Use antibiotics only for bacterial infections
- Identify the causative organism if possible
- Use the right antibiotic; do not rely on broad-range antibiotics
- Not stop antibiotics as soon as symptoms improve; finish the full course
- Not use antibiotics for most colds, coughs, bronchitis, sinus infections, and eye infections, which are caused by viruses.
It is argued that government legislation will aid in educating the public on the importance of restrictive use of antibiotics, not only for human clinical use but also for treating animals raised for human consumption.
Causes and risk factors
Schematic representation of how antibiotic resistance evolves via natural selection. The top section represents a population of bacteria before exposure to an antibiotic. The middle section shows the population directly after exposure, the phase in which selection took place. The last section shows the distribution of resistance in a new generation of bacteria. The legend indicates the resistance levels of individuals.
Antibiotic resistance can be a result of horizontal gene transfer, and also of unlinked point mutations in the pathogen genome and a rate of about 1 in 108 per chromosomal replication. The antibiotic action against the pathogen can be seen as an environmental pressure; those bacteria which have a mutation allowing them to survive will live on to reproduce. They will then pass this trait to their offspring, which will result in a fully resistant colony.
Several studies have demonstrated that patterns of antibiotic usage greatly affect the number of resistant organisms which develop. Overuse of broad-spectrum antibiotics, such as second- and third-generation cephalosporins, greatly hastens the development of methicillin resistance. Other factors contributing towards resistance include incorrect diagnosis, unnecessary prescriptions, improper use of antibiotics by patients, the impregnation of household items and children’s toys with low levels of antibiotics, and the administration of antibiotics by mouth in livestock for growth promotion. Also unsound practices in the pharmaceutical manufacturing industry can contribute towards the likeliness of creation antibiotic resistant strains. Researchers have recently demonstrated the bacterial protein LexA may play a key role in the acquisition of bacterial mutations.
Drug resistance occurs in several classes of pathogens:
- bacteria—antibiotic resistance
- viruses—resistance to antiviral drugs
- cancer cells
The four main mechanisms by which microorganisms exhibit resistance to antimicrobials are:
- Drug inactivation or modification: e.g. enzymatic deactivation of Penicillin G in some penicillin-resistant bacteria through the production of ?-lactamases. Antibiotic modification is the best known: the resistant bacteria retain the same sensitive target as antibiotic sensitive strains, but the antibiotic is prevented from reaching it. This happens, for example, with lactamases the lactamase enzymatically cleaves the four membered lactam ring, rendering the antibiotic inactive. Over 200 types of lactamase have been described (table). Most lactamases act to some degree against both penicillins and cephalosporins; others are more specific namely, cephalosporinases (for example, AmpC enzyme found in Enterobacter spp) or penicillinases (for example, Staphylococcus aureus penicillinase). Lactamases are widespread among many bacterial species (both Gram positive and Gram negative) and exhibit varying degrees of inhibition by lactamase inhibitors, such as clavulanic acid.
- Alterations in the primary site of action may mean that the antibiotic penetrates the cell and reaches the target site but is unable to inhibit the activity of the target because of structural changes in the molecule. Enterococci are regarded as being inherently resistant to cephalosporins because the enzymes responsible for cell wall synthesis (production of the polymer peptidoglycan) known as penicillin binding proteins have a low affinity for them and therefore are not inhibited. Most strains of Streptococcus pneumoniae are highly susceptible to both penicillins and cephalosporins but can acquire DNA from other bacteria, which changes the enzyme so that they develop a low affinity for penicillins and hence become resistant to inhibition by penicillins.3 The altered enzyme still synthesises peptidoglycan but it now has a different structure.4 Mutants of Streptococcus pyogenes that are resistant to penicillin and express altered penicillin binding proteins can be selected in the laboratory, but they have not been seen in patients, possibly because the cell wall can no longer bind the anti-phagocytic M protein.
- Alteration of metabolic pathway: e.g. some sulfonamide-resistant bacteria do not require para-aminobenzoic acid (PABA), an important precursor for the synthesis of folic acid and nucleic acids in bacteria inhibited by sulfonamides. Instead, like mammalian cells, they turn to utilizing preformed folic acid.
- Quick Efflux: Active efflux is a mechanism responsible for extrusion of toxic substances and antibiotics outside the cell, this is considered to be a vital part of xenobiotic metabolism. This mechanism is important in medicine as it can contribute to bacterial antibiotic resistance.Efflux systems function via an energy-dependent mechanism (Active transport) to pump out unwanted toxic substances through specific efflux pumps. Some efflux systems are drug-specific while others may accommodate multiple drugs, and thus contribute to bacterial multidrug resistance (MDR).
There are three known mechanisms of fluoroquinolone resistance. Some types of efflux pumps can act to decrease intracellular quinolone concentration. In gram-negative bacteria, plasmid-mediated resistance genes produce proteins that can bind to DNA gyrase, protecting it from the action of quinolones. Finally, mutations at key sites in DNA gyrase or Topoisomerase IV can decrease their binding affinity to quinolones, decreasing the drug’s effectiveness.
Bacterial efflux pumps are proteinaceous transporters localized in the cytoplasmic membrane of all kinds of cells. They are active transporters meaning that they require a source of chemical energy to perform their function. Some are primary active transporters utilizing Adenosine triphosphate hydrolysis as a source of energy, while others are secondary active transporters (uniporters, symporters or antiporters) in which transport is coupled to an electrochemical potential difference created by pumping out hydrogen or sodium ions outside the cell.Bacterial efflux transporters are classified into five major superfamilies, based on the amino acid sequence and the energy source used to export their substrates:
- The major facilitator superfamily (MFS);
- The ATP-binding cassette superfamily (ABC);
- The small multidrug resistance family (SMR);
- The resistance-nodulation-cell division superfamily (RND); and
- The Multi antimicrobial extrusion protein family (MATE).
Of these only the ABC superfamily are primary transporters, the rest being secondary transporters utilizing proton or sodium gradient as a source of energy. While MFS dominates in Gram positive bacteria , the RND family is unique to Gram-negatives.
In the case of imipenem resistant Pseudomonas aeruginosa, lack of the specific D2 porin confers resistance, as imipenem cannot penetrate the cell. This mechanism is also seen with low level resistance to fluoroquinolones and aminoglycosides. Increased efflux via an energy-requiring transport pump is a well recognised mechanism for resistance to tetracyclines and is encoded by a wide range of related genes, such as tet(A), that have become distributed in the enterobacteriaceae.
Although antibiotics are the most clinically important substrates of efflux systems, it is probable that most efflux pumps have other natural physiological functions. Examples include:
- The E.coli AcrAB efflux system which has a physiologic role of pumping out bile acids and fatty acids to lower their toxicity.
- The MFS family Ptr pump in Streptomyces pristinaespiralis appears to be an autoimmunity pump for this organism when it turns on production of pristinamycins I and II.
- The AcrAB–TolC system in E.coli is suspected to have a role in the transport of the calcium-channel components in the E. coli membrane.
- The MtrCDE system plays a protective role by providing resistance to faecal lipids in rectal isolates of Neisseria gonorrhoeae.
- The AcrAB efflux system of Erwinia amylovora is important for this organism’s virulence, plant (host) colonization and resistance to plant toxins.
The ability of efflux systems to recognize a large number of compounds other than their natural substrates is probably because substrate recognition is based on physicochemical properties, such as hydrophobicity, aromaticity and ionizable character rather than on defined chemical properties, as in classical enzyme-substrate or ligand-receptor recognition. Because most antibiotics are amphiphilic molecules – possessing both hydrophilic and hydrophobic characters, they are easily recognized by many efflux pumps.
Impact on antimicrobial resistance
The impact of efflux mechanisms on antimicrobial resistance is large, this is usually attributed to the following:
- The genetic elements encoding efflux pumps may be encoded on chromosomes and/or plasmids, thus contributing to both intrinsic (natural) and acquired resistance respectively. As an intrinsic mechanism of resistance, efflux pump genes can survive a hostile environment ( for example in the presence of antibiotics) which allows for the selection of mutants that over-express these genes. Being located on transpoable genetic elements as plasmids or transposons is also advantageous for the microorganisms as it allows for the easy spread of efflux genes between distant species.
- Antibiotics can act as inducers and regulators of the expression of some efflux pumps.
- Expression of several efflux pumps in a given bacterial species may lead to a broad spectrum of resistance when considering the shared substrates of some multi-drug efflux pumps, where one efflux pump may confer resistance to a wide range of antimicrobials.
Molecular epidemiology of resistance genes
Resistance in bacteria can be intrinsic or acquired. Intrinsic resistance is a naturally occurring trait arising from the biology of the organism for example, vancomycin resistance in Escherichia coli. Acquired resistance occurs when a bacterium that has been sensitive to antibiotics develops resistance this may happen by mutation or by acquisition of new DNA.
Mutation is a spontaneous event that occurs regardless of whether antibiotic is present. A bacterium carrying such a mutation is at a huge advantage as the susceptible cells are rapidly killed by the antibiotic, leaving a resistant subpopulation. Transferable resistance was recognised in 1959, when resistance genes found in shigella transferred to E coli via plasmids. Plasmids are self replicating circular pieces of DNA, smaller than the bacterial genome, which encode their transfer by replication into another bacterial strain or species. They can carry and transfer multiple resistance genes, which may be located on a section of DNA capable of transfer from one plasmid to another or to the genome a transposon (or “jumping gene”). Because the range of bacteria to which plasmids can spread is often limited, transposons are important in spreading resistance genes across such boundaries. The mecA gene found in MRSA may well have been acquired by transposition.7 Plasmid evolution can be complex, but modern molecular techniques can give an understanding (as is the case with the plasmids that contain the tetM gene and are found throughout the world in Neisseria gonorrhoeae).8
Bacteriophages (viruses that infect bacteria) can also transfer resistance, and this is frequently seen in staphylococci. When bacteria die they release DNA, which can be taken up by competent bacteria a process known as transformation. This process is increasingly recognised as important in the environment and is probably the main route for the spread of penicillin resistance in Streptococcus pneumoniae, by creation of “mosaic penicillin binding protein genes.
Origins of resistance genes
The origins of antibiotic resistance genes are obscure because at the time that antibiotics were introduced the biochemical and molecular basis of resistance was yet to be discovered. Bacteria collected between 1914 and 1950 (the Murray collection) were later found to be completely sensitive to antibiotics. They did, however, contain a range of plasmids capable of conjugative transfer.9 None of the Murray strains was resistant to sulphonamides, although these had been introduced in the mid-1930s; resistance was reported in the early 1940s in streptococci and gonococci.10 The introduction of streptomycin for treating tuberculosis was thwarted by the rapid development of resistance by mutation of the target genes. Mutation is now recognised as the commonest mechanism of resistance development in Mycobacterium tuberculosis, and the molecular nature of the mutations conferring resistance to most antituberculosis drugs is now known.11 Favourable mutations that arise in bacteria can be mobilised via insertion sequences and transposons on to plasmids and then transferred to different bacterial species.
In considering the evolution and dissemination of antibiotic resistance genes it is important to appreciate the rapidity of bacterial multiplication and the continual exchange of bacteria among animal, human, and agricultural hosts throughout the world. There is support for the notion that determinants of antibiotic resistance were not derived from the currently observed bacterial host in which the resistance plasmid is seen. DNA sequencing studies of lactamases and aminoglycoside inactivating enzymes show that despite similarities within the protein studies of the two families, there are substantial sequence differences. 12 13 As the evolutionary time frame has to be less than 50 years it is not possible to derive a model in which evolution could have occurred by mutation alone from common ancestral genes. They must have been derived from a large and diverse gene pool presumably already occurring in environmental bacteria. Many bacteria and fungi that produce antibiotics possess resistance determinants that are similar to those found in clinical bacteria.10 Gene exchange might occur in soil or, more likely, in the gut of humans or animals. It has been discovered that commercial antibiotic preparations contain DNA from the producing organism, and antibiotic resistance gene sequences can be identified by the polymerase chain reaction.14
Genes either exist in nature already or can emerge by mutation rapidly. Rapid mutation has been seen with (a) the TEM lactamase, resulting in an extension of the substrate profile to include third generation cephalosporins (first reported in Athens in 1963, one year after the introduction of ampicillin) and (b) the IMI-1 lactamase (reported from a Californian hospital before imipenem was approved for use in the United States).15 The selection pressure is heavy, and injudicious use of antibiotics, largely in medical practice, is probably responsible although agricultural and veterinary use contributes to resistance in human pathogens. The addition of antibiotics to animal feed or water, either for growth promotion or, more significantly, for mass treatment or prophylaxis (or both treatment and prophylaxis) in factory farmed animals, is having an unquantified effect on resistance levels.16 Bacteria clearly have a wondrous array of biochemical and genetic systems for ensuring the evolution and dissemination of antibiotic resistance.
Resistance mechanism to some important antibiotics
1. ß-lactam resistance
ß-lactams belong to a family of antibiotics which is characterized by a ß-lactam ring. Penicillins, cephalosporins, clavams (or oxapenams), cephamycins and carbapenems are members of this family. The integrity of the ß-lactam ring is necessary for the activity which results in the inactivation of a set of transpeptidases that catalyze the final cross-linking reactions of peptidoglycan synthesis. Resistance to ß-lactams in clinical isolates is primarily due to the hydrolysis of the antibiotic by a ß-lactamase. Mutational events resulting in the modification of PBPs (penicillin binding proteins) or cellular permeability can also lead to ß-lactam resistance. ß-lactamases constitute a heterogenous group of enzymes. Several classification schemes have been proposed according to their hydrolytic spectrum, susceptibility to inhibitors, genetic localisation (plasmidic or chromosomal), gene or amino-acid protein sequence. The functional classification scheme of ß-lactamases proposed by Bush, Jacoby and Medeiros (1995) defines four groups according to their substrate and inhibitor profiles. Group 1 are cephalosporinases that are not well inhibited by clavulanic acid; group 2 penicillinases, cephalosporinases, and broad-spectrum ß-lactamases that are generally inhibited by active site-directed ß-lactamase inhibitors; group 3 metallo-ß-lactamases that hydrolyze penicillins, cephalosporins, and carbapenems and that are poorly inhibited by almost all ß-lactam-containing molecules; group 4 penicillinases that are not well inhibited by clavulanic acid. Subgroups were also defined according to rates of hydrolysis of carbenicillin or cloxacillin (oxacillin) by group 2 penicillinases. The classification initially introduced by Ambler (1980) and based on the amino-acid sequence recognizes four molecular classes designated A to D. Classes A, C, and D gather evolutionarily distinct groups of serine enzymes, and class B the zinc-dependent (“EDTA-inhibited”) enzymes. Fig : ß-lactamases
Commonly used B-lactam resistance markers in molecular biology
The bla gene encoding the TEM-1 ß-lactamase is the most encountered AmpR marker used in molecular biology (pBR and pUC plasmids). TEM-1 is a widespread plasmidic ß-lactamase that attacks narrow-spectrum cephalosporins, cefamandole, and cefoperazone and all the anti-gram-negative-bacterium penicillins except temocillin. Aminothiazol chephalosporins, cephamycins, monobactams and carbapenems are resistant to its action. It belongs to the Bush-Jacoby-Medeiros group 2b and the molecular class A. The TEM-1 enzyme was first reported from an E. coli isolate in 1965 and is now the commonest ß-lactamase found in enterobacteriaceae. Resistance in more than 50% of AmpR E. coli clinical isolates is due to TEM-1. Most extended-spectrum ß-lactamases (ESBLs) derive from TEM-1, TEM-2 and SHV-1 by mutations generating 1- to 4-amino-acid sequence substitutions.
2. Aminoglycoside resistance
Aminoglycosides (Streptomycin, kanamycin, tobramycin, amikacin,…) are compounds that are characterized by the presense of an aminocyclitol ring linked to aminosugars in their structure. Their bactericidal activity is attributed to the irreversible binding to the ribosomes although their interaction with other cellular structures and metabolic processes has also been considered. They have a broad antimicrobial spectrum. They are active against aerobic and facultative aerobic Gram-negative bacilli and some Gram-positive bacteria of which staphylococci. Aminoglycosides are not active against anaerobes and rikettsia. Spectinomycin which is an aminocyclitol devoided of aminosugars is by extension included in the familiy of aminoglycosides. It also differs from them by its bacteriostatic ativity and by its way of action. Spectinomycin acts on protein synthesis during the mRNA-ribosome interaction and it does not lead to mistranslation like aminoglycosides do. Three mechanisms of resistance have been recognized, namely ribosome alteration, decreased permeability, and inactivation of the drugs by aminoglycoside modifying enzymes. The latter mechanism is of most clinical importance since the genes encoding aminoglycoside modifying enzymes can be disseminated by plasmids or transposons.
High level resistance to streptomycin and spectinomycin can result from single step mutations in chromosomal genes encoding ribosomal proteins: rpsL (or strA), rpsD (or ramA or sud2), rpsE (eps or spc or spcA). Mutations in strC (or strB) generate a low-level streptomycin resistance.
Absence of or alteration in the aminoglycoside transport system, inadequate membrane potential, modification in the LPS (lipopolysacchaccarides) phenotype can result in a cross resistance to all aminoglycosides.
Inactivation of aminoglycosides
These enzymes are classified into three major classes according to the type modification: AAC (acetyltransferases), ANT (nucleotidyltransferases or adenyltransferases), APH (phosphotransferases). This classification was extensively reviewed by Shaw et al. (1993).
Commonly used aminoglycoside resistance markers in molecular biology
ant(3”)-Ia (synonyms: aadA, aad(3”)(9))confers resistance to streptomycin and spectinomycin. The gene has been found in association with several transposons (Tn7, Tn21, …) and is ubiquitous among gram-negative bacteria.aph(3′)-II (synonyms: aphA-2, nptII) confers resistance to Km (Kanamycin), Neo (Neomycin), Prm (Paromomycin), Rsm (Ribostamycin), But (Butirosin), GmB (GentamycinB). This gene is rarely found in clinical isolates. aph(3′)-II is associated with transposon Tn5 and observed in gram-negative bacteria and Pseudomonas sp. However, its relative abundance in environmental KanR isolates seems to be low (Recorbet et al., 1992; Leff et al., 1993; Smalla et al., 1993).aph(3′)-III (synonyms: nptIII) confers resistance to Km (Kanamycin), Neo (Neomycin), Prm (Paromomycin), Rsm (Ribostamycin), Lvdm (Lividomycin), But (Butirosin), GmB (GentamycinB). Amk (Amikacin) and Isp (Isepamicin) are also modified in vitro, but according to the susceptibility standards established by NCCLS resistance is only expressed at a low level by many strains. aph(3′)-III is commonly distributed among gram-positive bacteria but has also been observed in Campylobacter spp.
nptIII is not frequent in molecular biology but can be found on some Agrobacterium vectors for plant transformation (Bevan, 1984).
3. Tetracycline resistance
Tetracyclines (tetracycline, doxycycline, minocycline, oxtetracycline) are antibiotics which inhibit the bacterial growth by stopping protein synthesis. They have been widely used for the past forty years as therapeutic agent in human and veterinary medicine but also as growth promotor in animal husbandry. The emergence of bacterial resistances to these antibiotics has nowadays limited their use. Three different specific mechanisms of tetracycline resistance have been identified so far: tetracycline efflux, ribosome protection and tetracycline modification.
Tetracycline efflux is achieved by an export protein from the major facilitator superfamily (MFS). The export protein was shown to function as an electroneutral antiport system which catalyzes the exchange of tetracycline-divalent-metal-cation complex for a proton. In Gram-negative bacteria the export protein contains 12 TMS (transmembrane fragments) whereas in Gram-positive bacteria it displays 14 TMS. Ribosome protection is mediated by a soluble protein which shares homolgy with the GTPases participating in protein synthesis, namely EF-Tu and EF-G. The third mechanism involves a cytoplasmic protein that chemically modifies tetracycline. This reaction takes only place in the presence of oxygen and NADPH and does not function in the natural host (Bacteroides). The two first mechanisms are the most widespread and most of their genes are normally acquired via transferable plasmids and/or transposons. These two mechanisms were observed both in aerobic and anaerobic Gram-negative or Gram-positive bacteria demonstrating their wide distribution among the bacterial kingdom. To date, about sixty-one tetracycline resistance genes have been sequenced and thirty-two classes of genes identified in non-producers and producers (Streptomyces). Each new class is identified by its inability to hybridize with any of the known tet genes under stringent conditions. A new nomenclature for the resistance determinants has been proposed for the future with the S. B. Levy group to coordinate the naming of the
Commonly used tetracycline resistance markers in molecular biology
Several tetracycline resistance determinants are currently used in molecular biology. The most encountered are the tetA genes of classes A (RP1, RP4 or Tn1721 derivatives), B (Tn10 derivatives) and C (pSC101 or pBR322 derivatives) encoding a tetracycline efflux system.These genes are regulated by a repressor protein (TetR). This feature has also been exploited to construct tightly regulated, high level mammalian expression systems by using the regulatory elements of the Tn10 tetracycline operon (Tet-OffTM and Tet-OnTM Expression Systems & Cell Lines,Clontech).The tetM gene from Tn916 which can be expressed both in Gram-positive and Gram-negative bacteria is also frequently used. Several Bacteroides/Escherichia shuttle vectors contain the tetQ gene. tetM and tetQ encode a soluble protein protecting the ribosome from the inhibiting effects of tetracycline. The distribution of these genes is given in the pages relating to the determinant classification.
Some Resistant pathogens
Staphylococcus aureus (colloquially known as “Staph aureus” or a Staph infection) is one of the major resistant pathogens. Found on the mucous membranes and the skin of around a third of the population, it is extremely adaptable to antibiotic pressure. It was the first bacterium in which penicillin resistance was found—in 1947, just four years after the drug started being mass-produced. Methicillin was then the antibiotic of choice, but has since been replaced by oxacillin due to significant kidney toxicity. MRSA (methicillin-resistant Staphylococcus aureus) was first detected in Britain in 1961 and is now “quite common” in hospitals. MRSA was responsible for 37% of fatal cases of blood poisoning in the UK in 1999, up from 4% in 1991. Half of all S. aureus infections in the US are resistant to penicillin, methicillin, tetracycline and erythromycin.
Methicillin Resistant Staphylococcus Aureus (MRSA) is acknowledged to be a human commensal and pathogen. MRSA has been found in cats, dogs and horses, where it can cause the same problems as it does in humans. Owners can transfer the organism to their pets and vice-versa, and MRSA in animals is generally believed to be derived from humans.
This left vancomycin as the only effective agent available at the time. However, strains with intermediate (4-8 ug/ml) levels of resistance, termed GISA (glycopeptide intermediate Staphylococcus aureus) or VISA (vancomycin intermediate Staphylococcus aureus), began appearing in the late 1990s. The first identified case was in Japan in 1996, and strains have since been found in hospitals in England, France and the US. The first documented strain with complete (>16 ug/ml) resistance to vancomycin, termed VRSA (Vancomycin-resistant Staphylococcus aureus) appeared in the United States in 2002.
A new class of antibiotics, oxazolidinones, became available in the 1990s, and the first commercially available oxazolidinone, linezolid, is comparable to vancomycin in effectiveness against MRSA. Linezolid-resistance in Staphylococcus aureus was reported in 2003.
CA-MRSA (Community-acquired MRSA) has now emerged as an epidemic that is responsible for rapidly progressive, fatal diseases including necrotizing pneumonia, severe sepsis and necrotizing fasciitis. Methicillin-resistant Staphylococcus aureus (MRSA) is the most frequently identified antimicrobial drug-resistant pathogen in US hospitals. The epidemiology of infections caused by MRSA is rapidly changing. In the past 10 years, infections caused by this organism have emerged in the community. The 2 MRSA clones in the United States most closely associated with community outbreaks, USA400 (MW2 strain, ST1 lineage) and USA300, often contain Panton-Valentine leukocidin (PVL) genes and, more frequently, have been associated with skin and soft tissue infections. Outbreaks of community-associated (CA)-MRSA infections have been reported in correctional facilities, among athletic teams, among military recruits, in newborn nurseries, and among active homosexual men. CA-MRSA infections now appear to be endemic in many urban regions and cause most CA-S. aureus infections.
Streptococcus and Enterococcus
Streptococcus pyogenes (Group A Streptococcus: GAS) infections can usually be treated with many different antibiotics. Early treatment may reduce the risk of death from invasive group A streptococcal disease. However, even the best medical care does not prevent death in every case. For those with very severe illness, supportive care in an intensive care unit may be needed. For persons with necrotizing fasciitis, surgery often is needed to remove damaged tissue. Strains of S. pyogenes resistant to macrolide antibiotics have emerged, however all strains remain uniformly sensitive to penicillin.
Resistance of Streptococcus pneumoniae to penicillin and other beta-lactams is increasing worldwide. The major mechanism of resistance involves the introduction of mutations in genes encoding penicillin-binding proteins. Selective pressure is thought to play an important role, and use of beta-lactam antibiotics has been implicated as a risk factor for infection and colonization. Streptococcus pneumoniae is responsible for pneumonia, bacteremia, otitis media, meningitis, sinusitis, peritonitis and arthritis.
Penicillin-resistant pneumonia caused by Streptococcus pneumoniae (commonly known as pneumococcus), was first detected in 1967, as was penicillin-resistant gonorrhea. Resistance to penicillin substitutes is also known as beyond S. aureus. By 1993 Escherichia coli was resistant to five fluoroquinolone variants. Mycobacterium tuberculosis is commonly resistant to isoniazid and rifampin and sometimes universally resistant to the common treatments. Other pathogens showing some resistance include Salmonella, Campylobacter, and Streptococci.
Enterococcus faecium is another superbug found in hospitals. Penicillin-Resistant Enterococcus was seen in 1983, vancomycin-resistant enterococcus (VRE) in 1987, and Linezolid-Resistant Enterococcus (LRE) in the late 1990s.
Pseudomonas aeruginosa is a highly prelevant opportunistic pathogen. One of the most worrisome characteristics of P. aeruginosa consists in its low antibiotic susceptibility. This low susceptibility is attributable to a concerted action of multidrug efflux pumps with chromosomally-encoded antibiotic resistance genes (e.g. mexAB-oprM, mexXY etc) and the low permeability of the bacterial cellular envelopes. Besides intrinsic resistance, P. aeruginosa easily develop acquired resistance either by mutation in chromosomally-encoded genes, or by the horizontal gene transfer of antibiotic resistance determinants. Development of multidrug resistance by P. aeruginosa isolates requires several different genetic events that include acquisition of different mutations and/or horizontal transfer of antibiotic resistance genes. Hypermutation favours the selection of mutation-driven antibiotic resistance in P. aeruginosa strains producing chronic infections, whereas the clustering of several different antibiotic resistance genes in integrons favours the concerted acquisition of antibiotic resistance determinants. Some recent studies have shown that phenotypic resistance associated to biofilm formation or to the emergence of small-colony-variants may be important in the response of P. aeruginosa populations to antibiotics treatment.
Clostridium difficile is a nosocomial pathogen that causes diarrheal disease in hospitals worldwide. Clindamycin-resistant C. difficile was reported as the causative agent of large outbreaks of diarrheal disease in hospitals in New York, Arizona, Florida and Massachusetts between 1989 and 1992. Geographically dispersed outbreaks of C. difficile strains resistant to fluoroquinolone antibiotics, such as Cipro (ciprofloxacin) and Levaquin (levofloxacin), were also reported in North America in 2005.
Salmonella and E. coli
E. coli and Salmonella come directly from contaminated food. Of the meat that is contaminated with E. coli, eighty percent of the bacteria are resistant to one or more drugs made; it causes bladder infections that are resistant to antibiotics (“HSUS Fact Sheet”). Salmonella was first found in humans in the 1970s and in some cases is resistant to as many as nine different antibiotics (“HSUS Fact Sheet”). When both bacterium are spread, serious health conditions arise. Many people are hospitalized each year after becoming infected, and some die as a result.
On the 5th November 2004, the Centers for Disease Control and Prevention (CDC) reported an increasing number of Acinetobacter baumannii bloodstream infections in patients at military medical facilities in which service members injured in the Iraq/Kuwait region during Operation Iraqi Freedom and in Afghanistan during Operation Enduring Freedom were treated. Most of these showed multidrug resistance (MRAB), with a few isolates resistant to all drugs tested.
We frequently refer to bacteria as being resistant to antibiotics, but rarely do we consider what that means. Even the most resistant bacterium can be inhibited or killed by a sufficiently high concentration of antibiotic; patients, however, would not be able to tolerate the high concentration required in some cases. Bacterial species vary tremendously in their susceptibility to an antibiotic for example, most strains of Streptococcus pneumoniae in Britain are inhibited by 0.01 mg/l of benzyl penicillin (the minimum inhibitory concentration), whereas for Escherichia coli 32-64 mg/l are required to inhibit growth, a level which cannot be achieved in the human body. This introduces the concept of clinical resistance, which is dependent on outcome and is all too often ignored. Clinical resistance is a complex concept in which the type of infecting bacterium, its location in the body, the distribution of the antibiotic in the body and its concentration at the site of infection, and the immune status of the patient all interact.
- Antibiotic resistance should be defined in terms of clinical outcomes, not laboratory methods
- Resistance occurs by means of four main mechanisms more than one may be present in a single bacterium
- Resistance mechanisms have probably evolved from genes present in organisms producing antibiotics
- Resistance genes occur not only in bacteria that carry disease but also in commensal bacteria, to which we are continuously exposed and which are found in food, the environment, and animals
- The plethora of genetic mechanisms for evolution and reassortment of antibiotic resistance genes ensures that useful genes will be disseminated rapidly
- Action must be taken to slow the rate of evolution and spread of antibiotic resistance genes, in which the biggest single factor is the amount of antibiotics used in human medicine and agriculture