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Recon methods.
Reconnaissance elements observing a potential ambush target on the move generally stayed 300–500 meters away. A "leapfrogging" recon technique can be used. Surveillance units were echeloned one behind the other. As the enemy drew close to the first, it fell back behind the last recon team, leaving an advance group in its place. This one in turn fell back as the enemy again closed the gap, and the cycle rotated. This method helped keep the enemy under continuous observation from a variety of vantage points, and allowed the recon groups to cover one another.
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Abzyme
An abzyme (from antibody and enzyme), also called catmab (from catalytic monoclonal antibody), and most often called catalytic antibody or sometimes catab, is a monoclonal antibody with catalytic activity. Abzymes are usually raised in lab animals immunized against synthetic haptens, but some natural abzymes can be found in normal humans (anti-vasoactive intestinal peptide autoantibodies) and in patients with autoimmune diseases such as systemic lupus erythematosus, where they can bind to and hydrolyze DNA. To date abzymes display only weak, modest catalytic activity and have not proved to be of any practical use. They are, however, subjects of considerable academic interest. Studying them has yielded important insights into reaction mechanisms, enzyme structure and function, catalysis, and the immune system itself.
Enzymes function by lowering the activation energy of the transition state of a chemical reaction, thereby enabling the formation of an otherwise less-favorable molecular intermediate between the reactant(s) and the product(s). If an antibody is developed to bind to a molecule that is structurally and electronically similar to the transition state of a given chemical reaction, the developed antibody will bind to, and stabilize, the transition state, just like a natural enzyme, lowering the activation energy of the reaction, and thus catalyzing the reaction. By raising an antibody to bind to a stable transition-state analog, a new and unique type of enzyme is produced.
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So far, all catalytic antibodies produced have displayed only modest, weak catalytic activity. The reasons for low catalytic activity for these molecules have been widely discussed. Possibilities indicate that factors beyond the binding site may play an important role, in particular through protein dynamics. Some abzymes have been engineered to use metal ions and other cofactors to improve their catalytic activity.
History.
The possibility of catalyzing a reaction by means of an antibody which binds the transition state was first suggested by William P. Jencks in 1969. In 1994 Peter G. Schultz and Richard A. Lerner received the prestigious Wolf Prize in Chemistry for developing catalytic antibodies for many reactions and popularizing their study into a significant sub-field of enzymology.
Abzymes in healthy human breast milk.
There are a broad range of abzymes in healthy human breast milk with DNAse, RNAse, and protease activity.
Potential HIV treatment.
In a June 2008 issue of the journal Autoimmunity Review, researchers S. Planque, Sudhir Paul, Ph.D., and Yasuhiro Nishiyama, Ph.D. of the University Of Texas Medical School at Houston announced that they have engineered an abzyme that degrades the superantigenic region of the gp120 CD4 binding site. This is the one part of the HIV virus outer coating that does not change, because it is the attachment point to T lymphocytes, the key cell in cell-mediated immunity. Once infected by HIV, patients produce antibodies to the more changeable parts of the viral coat. The antibodies are ineffective because of the virus' ability to change their coats rapidly. Because this protein gp120 is necessary for HIV to attach, it does not change across different strains and is a point of vulnerability across the entire range of the HIV variant population.
The abzyme does more than bind to the site: it catalytically destroys the site, rendering the virus inert, and then can attack other HIV viruses. A single abzyme molecule can destroy thousands of HIV viruses.
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Adaptive radiation
In evolutionary biology, adaptive radiation is a process in which organisms diversify rapidly from an ancestral species into a multitude of new forms, particularly when a change in the environment makes new resources available, alters biotic interactions or opens new environmental niches. Starting with a single ancestor, this process results in the speciation and phenotypic adaptation of an array of species exhibiting different morphological and physiological traits. The prototypical example of adaptive radiation is finch speciation on the Galapagos ("Darwin's finches"), but examples are known from around the world.
Characteristics.
Four features can be used to identify an adaptive radiation:
Conditions.
Adaptive radiations are thought to be triggered by an ecological opportunity or a new adaptive zone. Sources of ecological opportunity can be the loss of antagonists (competitors or predators), the evolution of a key innovation, or dispersal to a new environment. Any one of these ecological opportunities has the potential to result in an increase in population size and relaxed stabilizing (constraining) selection. As genetic diversity is positively correlated with population size the expanded population will have more genetic diversity compared to the ancestral population. With reduced stabilizing selection phenotypic diversity can also increase. In addition, intraspecific competition will increase, promoting divergent selection to use a wider range of resources. This ecological release provides the potential for ecological speciation and thus adaptive radiation.
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Occupying a new environment might take place under the following conditions:
Relationship between mass-extinctions and mass adaptive radiations.
A 2020 study found there to be no direct causal relationship between the proportionally most comparable mass radiations and extinctions in terms of "co-occurrence of species", substantially challenging the hypothesis of "creative mass extinctions".
Examples.
Darwin's finches.
Darwin's finches on the Galapagos Islands are a model system for the study of adaptive radiation. Today represented by approximately 15 species, Darwin's finches are Galapagos endemics famously adapted for a specialized feeding behavior (although one species, the Cocos finch ("Pinaroloxias inornata"), is not found in the Galapagos but on the island of Cocos south of Costa Rica). Darwin's finches are not actually finches in the true sense, but are members of the tanager family Thraupidae, and are derived from a single ancestor that arrived in the Galapagos from mainland South America perhaps just 3 million years ago.
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and are derived from a single ancestor that arrived in the Galapagos from mainland South America perhaps just 3 million years ago. Excluding the Cocos finch, each species of Darwin's finch is generally widely distributed in the Galapagos and fills the same niche on each island. For the ground finches, this niche is a diet of seeds, and they have thick bills to facilitate the consumption of these hard materials. The ground finches are further specialized to eat seeds of a particular size: the large ground finch ("Geospiza magnirostris") is the largest species of Darwin's finch and has the thickest beak for breaking open the toughest seeds, the small ground finch ("Geospiza fuliginosa") has a smaller beak for eating smaller seeds, and the medium ground finch ("Geospiza fortis") has a beak of intermediate size for optimal consumption of intermediately sized seeds (relative to "G. magnirostris" and "G. fuliginosa"). There is some overlap: for example, the most robust medium ground finches could have beaks larger than those of the smallest large ground finches.
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fuliginosa"). There is some overlap: for example, the most robust medium ground finches could have beaks larger than those of the smallest large ground finches. Because of this overlap, it can be difficult to tell the species apart by eye, though their songs differ. These three species often occur sympatrically, and during the rainy season in the Galapagos when food is plentiful, they specialize little and eat the same, easily accessible foods. It was not well-understood why their beaks were so adapted until Peter and Rosemary Grant studied their feeding behavior in the long dry season, and discovered that when food is scarce, the ground finches use their specialized beaks to eat the seeds that they are best suited to eat and thus avoid starvation.
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The other finches in the Galapagos are similarly uniquely adapted for their particular niche. The cactus finches ("Geospiza" sp.) have somewhat longer beaks than the ground finches that serve the dual purpose of allowing them to feed on Opuntia cactus nectar and pollen while these plants are flowering, but on seeds during the rest of the year. The warbler-finches ("Certhidea" sp.) have short, pointed beaks for eating insects. The woodpecker finch ("Camarhynchus pallidus") has a slender beak which it uses to pick at wood in search of insects; it also uses small sticks to reach insect prey inside the wood, making it one of the few animals that use tools.
The mechanism by which the finches initially diversified is still an area of active research. One proposition is that the finches were able to have a non-adaptive, allopatric speciation event on separate islands in the archipelago, such that when they reconverged on some islands, they were able to maintain reproductive isolation. Once they occurred in sympatry, niche specialization was favored so that the different species competed less directly for resources. This second, sympatric event was adaptive radiation.
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Cichlids of the African Great Lakes.
The haplochromine cichlid fishes in the Great Lakes of the East African Rift (particularly in Lake Tanganyika, Lake Malawi, and Lake Victoria) form the most speciose modern example of adaptive radiation. These lakes are believed to be home to about 2,000 different species of cichlid, spanning a wide range of ecological roles and morphological characteristics. Cichlids in these lakes fill nearly all of the roles typically filled by many fish families, including those of predators, scavengers, and herbivores, with varying dentitions and head shapes to match their dietary habits. In each case, the radiation events are only a few million years old, making the high level of speciation particularly remarkable. Several factors could be responsible for this diversity: the availability of a multitude of niches probably favored specialization, as few other fish taxa are present in the lakes (meaning that sympatric speciation was the most probable mechanism for initial specialization). Also, continual changes in the water level of the lakes during the Pleistocene (which often turned the largest lakes into several smaller ones) could have created the conditions for secondary allopatric speciation.
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Tanganyika cichlids.
Lake Tanganyika is the site from which nearly all the cichlid lineages of East Africa (including both riverine and lake species) originated. Thus, the species in the lake constitute a single adaptive radiation event but do not form a single monophyletic clade. Lake Tanganyika is also the least speciose of the three largest African Great Lakes, with only around 200 species of cichlid; however, these cichlids are more morphologically divergent and ecologically distinct than their counterparts in lakes Malawi and Victoria, an artifact of Lake Tanganyika's older cichlid fauna. Lake Tanganyika itself is believed to have formed 9–12 million years ago, putting a recent cap on the age of the lake's cichlid fauna. Many of Tanganyika's cichlids live very specialized lifestyles. The giant or emperor cichlid ("Boulengerochromis microlepis") is a piscivore often ranked the largest of all cichlids (though it competes for this title with South America's "Cichla temensis", the speckled peacock bass). It is thought that giant cichlids spawn only a single time, breeding in their third year and defending their young until they reach a large size, before dying of starvation some time thereafter.
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breeding in their third year and defending their young until they reach a large size, before dying of starvation some time thereafter. The three species of "Altolamprologus" are also piscivores, but with laterally compressed bodies and thick scales enabling them to chase prey into thin cracks in rocks without damaging their skin. "Plecodus straeleni" has evolved large, strangely curved teeth that are designed to scrape scales off of the sides of other fish, scales being its main source of food. "Gnathochromis permaxillaris" possesses a large mouth with a protruding upper lip, and feeds by opening this mouth downward onto the sandy lake bottom, sucking in small invertebrates. A number of Tanganyika's cichlids are shell-brooders, meaning that mating pairs lay and fertilize their eggs inside of empty shells on the lake bottom. "Lamprologus callipterus" is a unique egg-brooding species, with 15 cm-long males amassing collections of shells and guarding them in the hopes of attracting females (about 6 cm in length) to lay eggs in these shells.
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"Lamprologus callipterus" is a unique egg-brooding species, with 15 cm-long males amassing collections of shells and guarding them in the hopes of attracting females (about 6 cm in length) to lay eggs in these shells. These dominant males must defend their territories from three types of rival: (1) other dominant males looking to steal shells; (2) younger, "sneaker" males looking to fertilize eggs in a dominant male's territory; and (3) tiny, 2–4 cm "parasitic dwarf" males that also attempt to rush in and fertilize eggs in the dominant male's territory. These parasitic dwarf males never grow to the size of dominant males, and the male offspring of dominant and parasitic dwarf males grow with 100% fidelity into the form of their fathers. A number of other highly specialized Tanganyika cichlids exist aside from these examples, including those adapted for life in open lake water up to 200m deep.
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Malawi cichlids.
The cichlids of Lake Malawi constitute a "species flock" of up to 1000 endemic species. Only seven cichlid species in Lake Malawi are not a part of the species flock: the Eastern happy ("Astatotilapia calliptera"), the sungwa ("Serranochromis robustus"), and five tilapia species (genera "Oreochromis" and "Coptodon"). All of the other cichlid species in the lake are descendants of a single original colonist species, which itself was descended from Tanganyikan ancestors. The common ancestor of Malawi's species flock is believed to have reached the lake 3.4 million years ago at the earliest, making Malawi cichlids' diversification into their present numbers particularly rapid. Malawi's cichlids span a similarly range of feeding behaviors to those of Tanganyika, but also show signs of a much more recent origin. For example, all members of the Malawi species flock are mouth-brooders, meaning the female keeps her eggs in her mouth until they hatch; in almost all species, the eggs are also fertilized in the female's mouth, and in a few species, the females continue to guard their fry in their mouth after they hatch. Males of most species display predominantly blue coloration when mating. However, a number of particularly divergent species are known from Malawi, including the piscivorous "Nimbochromis livingtonii", which lies on its side in the substrate until small cichlids, perhaps drawn to its broken white patterning, come to inspect the predator - at which point they are swiftly eaten.
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Victoria's cichlids.
Lake Victoria's cichlids are also a species flock, once composed of some 500 or more species. The deliberate introduction of the Nile Perch ("Lates niloticus") in the 1950s proved disastrous for Victoria cichlids, and the collective biomass of the Victoria cichlid species flock has decreased substantially and an unknown number of species have become extinct. However, the original range of morphological and behavioral diversity seen in the lake's cichlid fauna is still mostly present today, if endangered. These again include cichlids specialized for niches across the trophic spectrum, as in Tanganyika and Malawi, but again, there are standouts. Victoria is famously home to many piscivorous cichlid species, some of which feed by sucking the contents out of mouthbrooding females' mouths. Victoria's cichlids constitute a far younger radiation than even that of Lake Malawi, with estimates of the age of the flock ranging from 200,000 years to as little as 14,000.
Adaptive radiation in Hawaii.
Hawaii has served as the site of a number of adaptive radiation events, owing to its isolation, recent origin, and large land area. The three most famous examples of these radiations are presented below, though insects like the Hawaiian drosophilid flies and "Hyposmocoma" moths have also undergone adaptive radiation.
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Hawaiian honeycreepers.
The Hawaiian honeycreepers form a large, highly morphologically diverse species group of birds that began radiating in the early days of the Hawaiian archipelago. While today only 17 species are known to persist in Hawaii (3 more may or may not be extinct), there were more than 50 species prior to Polynesian colonization of the archipelago (between 18 and 21 species have gone extinct since the discovery of the islands by westerners). The Hawaiian honeycreepers are known for their beaks, which are specialized to satisfy a wide range of dietary needs: for example, the beak of the ʻakiapōlāʻau ("Hemignathus wilsoni") is characterized by a short, sharp lower mandible for scraping bark off of trees, and the much longer, curved upper mandible is used to probe the wood underneath for insects. Meanwhile, the ʻiʻiwi ("Drepanis coccinea") has a very long curved beak for reaching nectar deep in "Lobelia" flowers. An entire clade of Hawaiian honeycreepers, the tribe Psittirostrini, is composed of thick-billed, mostly seed-eating birds, like the Laysan finch ("Telespiza cantans"). In at least some cases, similar morphologies and behaviors appear to have evolved convergently among the Hawaiian honeycreepers; for example, the short, pointed beaks of "Loxops" and "Oreomystis" evolved separately despite once forming the justification for lumping the two genera together. The Hawaiian honeycreepers are believed to have descended from a single common ancestor some 15 to 20 million years ago, though estimates range as low as 3.5 million years.
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Hawaiian silverswords.
Adaptive radiation is not a strictly vertebrate phenomenon, and examples are also known from among plants. The most famous example of adaptive radiation in plants is quite possibly the Hawaiian silverswords, named for alpine desert-dwelling "Argyroxiphium" species with long, silvery leaves that live for up to 20 years before growing a single flowering stalk and then dying. The Hawaiian silversword alliance consists of twenty-eight species of Hawaiian plants which, aside from the namesake silverswords, includes trees, shrubs, vines, cushion plants, and more. The silversword alliance is believed to have originated in Hawaii no more than 6 million years ago, making this one of Hawaii's youngest adaptive radiation events. This means that the silverswords evolved on Hawaii's modern high islands, and descended from a single common ancestor that arrived on Kauai from western North America. The closest modern relatives of the silverswords today are California tarweeds of the family Asteraceae.
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Hawaiian lobelioids.
Hawaii is also the site of a separate major floral adaptive radiation event: the Hawaiian lobelioids. The Hawaiian lobelioids are significantly more speciose than the silverswords, perhaps because they have been present in Hawaii for so much longer: they descended from a single common ancestor who arrived in the archipelago up to 15 million years ago. Today the Hawaiian lobelioids form a clade of over 125 species, including succulents, trees, shrubs, epiphytes, etc. Many species have been lost to extinction and many of the surviving species endangered.
Caribbean anoles.
Anole lizards are distributed broadly in the New World, from the Southeastern US to South America. With over 400 species currently recognized, often placed in a single genus ("Anolis"), they constitute one of the largest radiation events among all lizards. Anole radiation on the mainland has largely been a process of speciation, and is not adaptive to any great degree, but anoles on each of the Greater Antilles (Cuba, Hispaniola, Puerto Rico, and Jamaica) have adaptively radiated in separate, convergent ways.
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Anole radiation on the mainland has largely been a process of speciation, and is not adaptive to any great degree, but anoles on each of the Greater Antilles (Cuba, Hispaniola, Puerto Rico, and Jamaica) have adaptively radiated in separate, convergent ways. On each of these islands, anoles have evolved with such a consistent set of morphological adaptations that each species can be assigned to one of six "ecomorphs": trunk–ground, trunk–crown, grass–bush, crown–giant, twig, and trunk. Take for example crown–giants from each of these islands: the Cuban "Anolis luteogularis", Hispaniola's "Anolis ricordii", Puerto Rico's "Anolis cuvieri", and Jamaica's "Anolis garmani" (Cuba and Hispaniola are both home to more than one species of crown–giant). These anoles are all large, canopy-dwelling species with large heads and large lamellae (scales on the undersides of the fingers and toes that are important for traction in climbing), and yet none of these species are particularly closely related and appear to have evolved these similar traits independently. and yet none of these species are particularly closely related and appear to have evolved these similar traits independently. The same can be said of the other five ecomorphs across the Caribbean's four largest islands. Much like in the case of the cichlids of the three largest African Great Lakes, each of these islands is home to its own convergent "Anolis" adaptive radiation event.
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Other examples.
Presented above are the most well-documented examples of modern adaptive radiation, but other examples are known. Populations of three-spined sticklebacks have repeatedly diverged and evolved into distinct ecotypes. On Madagascar, birds of the family Vangidae are marked by very distinct beak shapes to suit their ecological roles. Madagascan mantellid frogs have radiated into forms that mirror other tropical frog faunas, with the brightly colored mantellas ("Mantella") having evolved convergently with the Neotropical poison dart frogs of Dendrobatidae, while the arboreal "Boophis" species are the Madagascan equivalent of tree frogs and glass frogs. The pseudoxyrhophiine snakes of Madagascar have evolved into fossorial, arboreal, terrestrial, and semi-aquatic forms that converge with the colubroid faunas in the rest of the world. These Madagascan examples are significantly older than most of the other examples presented here: Madagascar's fauna has been evolving in isolation since the island split from India some 88 million years ago, and the Mantellidae originated around 50 mya. Older examples are known: the K-Pg extinction event, which caused the disappearance of the dinosaurs and most other reptilian megafauna 65 million years ago, is seen as having triggered a global adaptive radiation event that created the mammal diversity that exists today. Also the Cambrian Explosion, where vacant niches left by the extinction of Ediacaran biota during End-Ediacaran mass extinction were filled up by the emergence of new phyla.
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Agarose gel electrophoresis
Agarose gel electrophoresis is a method of gel electrophoresis used in biochemistry, molecular biology, genetics, and clinical chemistry to separate a mixed population of macromolecules such as DNA or proteins in a matrix of agarose, one of the two main components of agar. The proteins may be separated by charge and/or size (isoelectric focusing agarose electrophoresis is essentially size independent), and the DNA and RNA fragments by length. Biomolecules are separated by applying an electric field to move the charged molecules through an agarose matrix, and the biomolecules are separated by size in the agarose gel matrix.
Agarose gel is easy to cast, has relatively fewer charged groups, and is particularly suitable for separating DNA of size range most often encountered in laboratories, which accounts for the popularity of its use. The separated DNA may be viewed with stain, most commonly under UV light, and the DNA fragments can be extracted from the gel with relative ease. Most agarose gels used are between 0.7–2% dissolved in a suitable electrophoresis buffer.
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Properties of agarose gel.
Agarose gel is a three-dimensional matrix formed of helical agarose molecules in supercoiled bundles that are aggregated into three-dimensional structures with channels and pores through which biomolecules can pass. The 3-D structure is held together with hydrogen bonds and can therefore be disrupted by heating back to a liquid state. The melting temperature is different from the gelling temperature, depending on the sources, agarose gel has a gelling temperature of and a melting temperature of . Low-melting and low-gelling agaroses made through chemical modifications are also available.
Agarose gel has large pore size and good gel strength, making it suitable as an anticonvection medium for the electrophoresis of DNA and large protein molecules. The pore size of a 1% gel has been estimated from 100 nm to 200–500 nm, and its gel strength allows gels as dilute as 0.15% to form a slab for gel electrophoresis. Low-concentration gels (0.1–0.2%) however are fragile and therefore hard to handle. Agarose gel has lower resolving power than polyacrylamide gel for DNA but has a greater range of separation, and is therefore used for DNA fragments of usually 50–20,000 bp in size. The limit of resolution for standard agarose gel electrophoresis is around 750 kb, but resolution of over 6 Mb is possible with pulsed field gel electrophoresis (PFGE). It can also be used to separate large proteins, and it is the preferred matrix for the gel electrophoresis of particles with effective radii larger than 5–10 nm. A 0.9% agarose gel has pores large enough for the entry of bacteriophage T4.
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The agarose polymer contains charged groups, in particular pyruvate and sulfate. These negatively charged groups create a flow of water in the opposite direction to the movement of DNA in a process called electroendosmosis (EEO), and can therefore retard the movement of DNA and cause blurring of bands. Higher concentration gels would have higher electroendosmotic flow. Low EEO agarose is therefore generally preferred for use in agarose gel electrophoresis of nucleic acids, but high EEO agarose may be used for other purposes. The lower sulfate content of low EEO agarose, particularly low-melting point (LMP) agarose, is also beneficial in cases where the DNA extracted from gel is to be used for further manipulation as the presence of contaminating sulfates may affect some subsequent procedures, such as ligation and PCR. Zero EEO agaroses however are undesirable for some applications as they may be made by adding positively charged groups and such groups can affect subsequent enzyme reactions. Electroendosmosis is a reason agarose is used in preference to agar as the agaropectin component in agar contains a significant amount of negatively charged sulfate and carboxyl groups. The removal of agaropectin in agarose substantially reduces the EEO, as well as reducing the non-specific adsorption of biomolecules to the gel matrix. However, for some applications such as the electrophoresis of serum proteins, a high EEO may be desirable, and agaropectin may be added in the gel used.
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Migration of nucleic acids in agarose gel.
Factors affecting migration of nucleic acid in gel.
A number of factors can affect the migration of nucleic acids: the dimension of the gel pores (gel concentration), size of DNA being electrophoresed, the voltage used, the ionic strength of the buffer, and the concentration of intercalating dye such as ethidium bromide if used during electrophoresis.
Smaller molecules travel faster than larger molecules in gel, and double-stranded DNA moves at a rate that is inversely proportional to the logarithm of the number of base pairs. This relationship however breaks down with very large DNA fragments, and separation of very large DNA fragments requires the use of pulsed field gel electrophoresis (PFGE), which applies alternating current from different directions and the large DNA fragments are separated as they reorient themselves with the changing field.
For standard agarose gel electrophoresis, larger molecules are resolved better using a low concentration gel while smaller molecules separate better at high concentration gel. Higher concentration gels, however, require longer run times (sometimes days).
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The movement of the DNA may be affected by the conformation of the DNA molecule, for example, supercoiled DNA usually moves faster than relaxed DNA because it is tightly coiled and hence more compact. In a normal plasmid DNA preparation, multiple forms of DNA may be present. Gel electrophoresis of the plasmids would normally show the negatively supercoiled form as the main band, while nicked DNA (open circular form) and the relaxed closed circular form appears as minor bands. The rate at which the various forms move however can change using different electrophoresis conditions, and the mobility of larger circular DNA may be more strongly affected than linear DNA by the pore size of the gel.
Ethidium bromide which intercalates into circular DNA can change the charge, length, as well as the superhelicity of the DNA molecule, therefore its presence in gel during electrophoresis can affect its movement. For example, the positive charge of ethidium bromide can reduce the DNA movement by 15%. Agarose gel electrophoresis can be used to resolve circular DNA with different supercoiling topology.
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DNA damage due to increased cross-linking will also reduce electrophoretic DNA migration in a dose-dependent way.
The rate of migration of the DNA is proportional to the voltage applied, i.e. the higher the voltage, the faster the DNA moves. The resolution of large DNA fragments however is lower at high voltage. The mobility of DNA may also change in an unsteady field – in a field that is periodically reversed, the mobility of DNA of a particular size may drop significantly at a particular cycling frequency. This phenomenon can result in band inversion in field inversion gel electrophoresis (FIGE), whereby larger DNA fragments move faster than smaller ones.
Mechanism of migration and separation.
The negative charge of its phosphate backbone moves the DNA towards the positively charged anode during electrophoresis. However, the migration of DNA molecules in solution, in the absence of a gel matrix, is independent of molecular weight during electrophoresis. The gel matrix is therefore responsible for the separation of DNA by size during electrophoresis, and a number of models exist to explain the mechanism of separation of biomolecules in gel matrix. A widely accepted one is the Ogston model which treats the polymer matrix as a sieve. A globular protein or a random coil DNA moves through the interconnected pores, and the movement of larger molecules is more likely to be impeded and slowed down by collisions with the gel matrix, and the molecules of different sizes can therefore be separated in this sieving process.
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The Ogston model however breaks down for large molecules whereby the pores are significantly smaller than size of the molecule. For DNA molecules of size greater than 1 kb, a reptation model (or its variants) is most commonly used. This model assumes that the DNA can crawl in a "snake-like" fashion (hence "reptation") through the pores as an elongated molecule. A biased reptation model applies at higher electric field strength, whereby the leading end of the molecule become strongly biased in the forward direction and pulls the rest of the molecule along. Real-time fluorescence microscopy of stained molecules, however, showed more subtle dynamics during electrophoresis, with the DNA showing considerable elasticity as it alternately stretching in the direction of the applied field and then contracting into a ball, or becoming hooked into a U-shape when it gets caught on the polymer fibres.
General procedure.
The details of an agarose gel electrophoresis experiment may vary depending on methods, but most follow a general procedure.
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Casting of gel.
The gel is prepared by dissolving the agarose powder in an appropriate buffer, such as TAE or TBE, to be used in electrophoresis. The agarose is dispersed in the buffer before heating it to near-boiling point, but avoid boiling. The melted agarose is allowed to cool sufficiently before pouring the solution into a cast as the cast may warp or crack if the agarose solution is too hot. A comb is placed in the cast to create wells for loading sample, and the gel should be completely set before use.
The concentration of gel affects the resolution of DNA separation. The agarose gel is composed of microscopic pores through which the molecules travel, and there is an inverse relationship between the pore size of the agarose gel and the concentration – pore size decreases as the density of agarose fibers increases. High gel concentration improves separation of smaller DNA molecules, while lowering gel concentration permits large DNA molecules to be separated. The process allows fragments ranging from 50 base pairs to several mega bases to be separated depending on the gel concentration used. The concentration is measured in weight of agarose over volume of buffer used (g/ml). For a standard agarose gel electrophoresis, a 0.8% gel gives good separation or resolution of large 5–10kb DNA fragments, while 2% gel gives good resolution for small 0.2–1kb fragments. 1% gels is often used for a standard electrophoresis. High percentage gels are often brittle and may not set evenly, while low percentage gels (0.1-0.2%) are fragile and not easy to handle. Low-melting-point (LMP) agarose gels are also more fragile than normal agarose gel. Low-melting point agarose may be used on its own or simultaneously with standard agarose for the separation and isolation of DNA. PFGE and FIGE are often done with high percentage agarose gels.
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Loading of samples.
Once the gel has set, the comb is removed, leaving wells where DNA samples can be loaded. Loading buffer is mixed with the DNA sample before the mixture is loaded into the wells. The loading buffer contains a dense compound, which may be glycerol, sucrose, or Ficoll, that raises the density of the sample so that the DNA sample may sink to the bottom of the well. If the DNA sample contains residual ethanol after its preparation, it may float out of the well. The loading buffer also includes colored dyes such as xylene cyanol and bromophenol blue used to monitor the progress of the electrophoresis. The DNA samples are loaded using a pipette.
Electrophoresis.
Agarose gel electrophoresis is most commonly done horizontally in a subaquaeous mode whereby the slab gel is completely submerged in buffer during electrophoresis. It is also possible, but less common, to perform the electrophoresis vertically, as well as horizontally with the gel raised on agarose legs using an appropriate apparatus. The buffer used in the gel is the same as the running buffer in the electrophoresis tank, which is why electrophoresis in the subaquaeous mode is possible with agarose gel.
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For optimal resolution of DNA greater than 2kb in size in standard gel electrophoresis, 5 to 8 V/cm is recommended (the distance in cm refers to the distance between electrodes, therefore this recommended voltage would be 5 to 8 multiplied by the distance between the electrodes in cm). Voltage may also be limited by the fact that it heats the gel and may cause the gel to melt if it is run at high voltage for a prolonged period, especially if the gel used is LMP agarose gel. Too high a voltage may also reduce resolution, as well as causing band streaking for large DNA molecules. Too low a voltage may lead to broadening of band for small DNA fragments due to dispersion and diffusion.
Since DNA is not visible in natural light, the progress of the electrophoresis is monitored using colored dyes. Xylene cyanol (light blue color) comigrates large DNA fragments, while Bromophenol blue (dark blue) comigrates with the smaller fragments. Less commonly used dyes include Cresol Red and Orange G which migrate ahead of bromophenol blue. A DNA marker is also run together for the estimation of the molecular weight of the DNA fragments. Note however that the size of a circular DNA like plasmids cannot be accurately gauged using standard markers unless it has been linearized by restriction digest, alternatively a supercoiled DNA marker may be used.
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Staining and visualization.
DNA as well as RNA are normally visualized by staining with ethidium bromide, which intercalates into the major grooves of the DNA and fluoresces under UV light. The intercalation depends on the concentration of DNA and thus, a band with high intensity will indicate a higher amount of DNA compared to a band of less intensity. The ethidium bromide may be added to the agarose solution before it gels, or the DNA gel may be stained later after electrophoresis. Destaining of the gel is not necessary but may produce better images. Other methods of staining are available; examples are MIDORI Green, SYBR Green, GelRed, methylene blue, brilliant cresyl blue, Nile blue sulfate, and crystal violet. SYBR Green, GelRed and other similar commercial products are sold as safer alternatives to ethidium bromide as it has been shown to be mutagenic in Ames test, although the carcinogenicity of ethidium bromide has not actually been established. SYBR Green requires the use of a blue-light transilluminator. DNA stained with crystal violet can be viewed under natural light without the use of a UV transilluminator which is an advantage, however it may not produce a strong band.
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When stained with ethidium bromide, the gel is viewed with an ultraviolet (UV) transilluminator. The UV light excites the electrons within the aromatic ring of ethidium bromide, and once they return to the ground state, light is released, making the DNA and ethidium bromide complex fluoresce. Standard transilluminators use wavelengths of 302/312-nm (UV-B), however exposure of DNA to UV radiation for as little as 45 seconds can produce damage to DNA and affect subsequent procedures, for example reducing the efficiency of transformation, "in vitro" transcription, and PCR. Exposure of DNA to UV radiation therefore should be limited. Using a higher wavelength of 365 nm (UV-A range) causes less damage to the DNA but also produces much weaker fluorescence with ethidium bromide. Where multiple wavelengths can be selected in the transilluminator, shorter wavelength can be used to capture images, while longer wavelength should be used if it is necessary to work on the gel for any extended period of time.
The transilluminator apparatus may also contain image capture devices, such as a digital or polaroid camera, that allow an image of the gel to be taken or printed.
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For gel electrophoresis of protein, the bands may be visualised with Coomassie or silver stains.
Downstream procedures.
The separated DNA bands are often used for further procedures, and a DNA band may be cut out of the gel as a slice, dissolved and purified. Contaminants however may affect some downstream procedures such as PCR, and low melting point agarose may be preferred in some cases as it contains fewer of the sulfates that can affect some enzymatic reactions. The gels may also be used for blotting techniques.
Buffers.
In general, the ideal buffer should have good conductivity, produce less heat and have a long life. There are a number of buffers used for agarose electrophoresis; common ones for nucleic acids include tris/acetate/EDTA (TAE) and tris/borate/EDTA (TBE). The buffers used contain EDTA to inactivate many nucleases which require divalent cation for their function. The borate in TBE buffer can be problematic as borate can polymerize, and/or interact with cis diols such as those found in RNA. TAE has the lowest buffering capacity, but it provides the best resolution for larger DNA. This means a lower voltage and more time, but a better product.
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Many other buffers have been proposed, e.g. lithium borate (LB), iso electric histidine, pK matched goods buffers, etc.; in most cases the purported rationale is lower current (less heat) and or matched ion mobilities, which leads to longer buffer life. Tris-phosphate buffer has high buffering capacity but cannot be used if DNA extracted is to be used in phosphate sensitive reaction. LB is relatively new and is ineffective in resolving fragments larger than 5 kbp; However, with its low conductivity, a much higher voltage could be used (up to 35 V/cm), which means a shorter analysis time for routine electrophoresis. As low as one base pair size difference could be resolved in 3% agarose gel with an extremely low conductivity medium (1 mM lithium borate).
Other buffering system may be used in specific applications, for example, barbituric acid-sodium barbiturate or tris-barbiturate buffers may be used for in agarose gel electrophoresis of proteins, for example in the detection of abnormal distribution of proteins.
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Applications.
Agarose gels are easily cast and handled compared to other matrices and nucleic acids are not chemically altered during electrophoresis. Samples are also easily recovered. After the experiment is finished, the resulting gel can be stored in a plastic bag in a refrigerator.
Electrophoresis is performed in buffer solutions to reduce pH changes due to the electric field, which is important because the charge of DNA and RNA depends on pH, but running for too long can exhaust the buffering capacity of the solution. Further, different preparations of genetic material may not migrate consistently with each other, for morphological or other reasons.
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Allele
An allele is a variant of the sequence of nucleotides at a particular location, or locus, on a DNA molecule.
Alleles can differ at a single position through single nucleotide polymorphisms (SNP), but they can also have insertions and deletions of up to several thousand base pairs.
Most alleles observed result in little or no change in the function or amount of the gene product(s) they code or regulate for. However, sometimes different alleles can result in different observable phenotypic traits, such as different pigmentation. A notable example of this is Gregor Mendel's discovery that the white and purple flower colors in pea plants were the result of a single gene with two alleles.
Nearly all multicellular organisms have two sets of chromosomes at some point in their biological life cycle; that is, they are diploid. For a given locus, if the two chromosomes contain the same allele, they, and the organism, are homozygous with respect to that allele. If the alleles are different, they, and the organism, are heterozygous with respect to those alleles.
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Popular definitions of 'allele' typically refer only to different alleles within genes. For example, the ABO blood grouping is controlled by the ABO gene, which has six common alleles (variants). In population genetics, nearly every living human's phenotype for the ABO gene is some combination of just these six alleles.
Etymology.
The word "allele" is a short form of "allelomorph" ("other form", a word coined by British geneticists William Bateson and Edith Rebecca Saunders) in the 1900s, which was used in the early days of genetics to describe variant forms of a gene detected in different phenotypes and identified to cause the differences between them. It derives from the Greek prefix ἀλληλο-, "allelo-", meaning "mutual", "reciprocal", or "each other", which itself is related to the Greek adjective ἄλλος, "allos" (cognate with Latin "alius"), meaning "other".
Alleles that lead to dominant or recessive phenotypes.
In many cases, genotypic interactions between the two alleles at a locus can be described as dominant or recessive, according to which of the two homozygous phenotypes the heterozygote most resembles. Where the heterozygote is indistinguishable from one of the homozygotes, the allele expressed is the one that leads to the "dominant" phenotype, and the other allele is said to be "recessive". The degree and pattern of dominance varies among loci. This type of interaction was first formally-described by Gregor Mendel. However, many traits defy this simple categorization and the phenotypes are modelled by co-dominance and polygenic inheritance.
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The term "wild type" allele is sometimes used to describe an allele that is thought to contribute to the typical phenotypic character as seen in "wild" populations of organisms, such as fruit flies ("Drosophila melanogaster"). Such a "wild type" allele was historically regarded as leading to a dominant (overpowering – always expressed), common, and normal phenotype, in contrast to "mutant" alleles that lead to recessive, rare, and frequently deleterious phenotypes. It was formerly thought that most individuals were homozygous for the "wild type" allele at most gene loci, and that any alternative "mutant" allele was found in homozygous form in a small minority of "affected" individuals, often as genetic diseases, and more frequently in heterozygous form in "carriers" for the mutant allele. It is now appreciated that most or all gene loci are highly polymorphic, with multiple alleles, whose frequencies vary from population to population, and that a great deal of genetic variation is hidden in the form of alleles that do not produce obvious phenotypic differences. Wild type alleles are often denoted by a superscript plus sign ("i.e.", "p" for an allele "p").
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Multiple alleles.
A population or species of organisms typically includes multiple alleles at each locus among various individuals. Allelic variation at a locus is measurable as the number of alleles (polymorphism) present, or the proportion of heterozygotes in the population. A null allele is a gene variant that lacks the gene's normal function because it either is not expressed, or the expressed protein is inactive.
For example, at the gene locus for the ABO blood type carbohydrate antigens in humans, classical genetics recognizes three alleles, IA, IB, and i, which determine compatibility of blood transfusions. Any individual has one of six possible genotypes (IAIA, IAi, IBIB, IBi, IAIB, and ii) which produce one of four possible phenotypes: "Type A" (produced by IAIA homozygous and IAi heterozygous genotypes), "Type B" (produced by IBIB homozygous and IBi heterozygous genotypes), "Type AB" produced by IAIB heterozygous genotype, and "Type O" produced by ii homozygous genotype. (It is now known that each of the A, B, and O alleles is actually a class of multiple alleles with different DNA sequences that produce proteins with identical properties: more than 70 alleles are known at the ABO locus. Hence an individual with "Type A" blood may be an AO heterozygote, an AA homozygote, or an AA heterozygote with two different "A" alleles.)
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Genotype frequencies.
The frequency of alleles in a diploid population can be used to predict the frequencies of the corresponding genotypes (see Hardy–Weinberg principle). For a simple model, with two alleles;
where "p" is the frequency of one allele and "q" is the frequency of the alternative allele, which necessarily sum to unity. Then, "p"2 is the fraction of the population homozygous for the first allele, 2"pq" is the fraction of heterozygotes, and "q"2 is the fraction homozygous for the alternative allele. If the first allele is dominant to the second then the fraction of the population that will show the dominant phenotype is "p"2 + 2"pq", and the fraction with the recessive phenotype is "q"2.
With three alleles:
In the case of multiple alleles at a diploid locus, the number of possible genotypes (G) with a number of alleles (a) is given by the expression:
Allelic dominance in genetic disorders.
A number of genetic disorders are caused when an individual inherits two recessive alleles for a single-gene trait. Recessive genetic disorders include albinism, cystic fibrosis, galactosemia, phenylketonuria (PKU), and Tay–Sachs disease. Other disorders are also due to recessive alleles, but because the gene locus is located on the X chromosome, so that males have only one copy (that is, they are hemizygous), they are more frequent in males than in females. Examples include red–green color blindness and fragile X syndrome.
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Other disorders, such as Huntington's disease, occur when an individual inherits only one dominant allele.
Epialleles.
While heritable traits are typically studied in terms of genetic alleles, epigenetic marks such as DNA methylation can be inherited at specific genomic regions in certain species, a process termed transgenerational epigenetic inheritance. The term "epiallele" is used to distinguish these heritable marks from traditional alleles, which are defined by nucleotide sequence. A specific class of epiallele, the metastable epialleles, has been discovered in mice and in humans which is characterized by stochastic (probabilistic) establishment of epigenetic state that can be mitotically inherited.
Idiomorph.
The term "idiomorph", from Greek 'morphos' (form) and 'idio' (singular, unique), was introduced in 1990 in place of "allele" to denote sequences at the same locus in different strains that have no sequence similarity and probably do not share a common phylogenetic relationship. It is used mainly in the genetic research of mycology.
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Ampicillin
Ampicillin is an antibiotic belonging to the aminopenicillin class of the penicillin family. The drug is used to prevent and treat several bacterial infections, such as respiratory tract infections, urinary tract infections, meningitis, salmonellosis, and endocarditis. It may also be used to prevent group B streptococcal infection in newborns. It is used by mouth, by injection into a muscle, or intravenously.
Common side effects include rash, nausea, and diarrhea. It should not be used in people who are allergic to penicillin. Serious side effects may include "Clostridioides difficile" colitis or anaphylaxis. While usable in those with kidney problems, the dose may need to be decreased. Its use during pregnancy and breastfeeding appears to be generally safe.
Ampicillin was discovered in 1958 and came into commercial use in 1961. It is on the World Health Organization's List of Essential Medicines. The World Health Organization classifies ampicillin as critically important for human medicine. It is available as a generic medication.
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Medical uses.
Diseases.
Ampicillin used to also be used to treat gonorrhea, but there are now too many strains resistant to penicillins.
Bacteria.
Ampicillin is used to treat infections by many gram-positive and gram-negative bacteria. It was the first "broad spectrum" penicillin with activity against gram-positive bacteria, including "Streptococcus pneumoniae", "Streptococcus pyogenes", some isolates of "Staphylococcus aureus" (but not penicillin-resistant or methicillin-resistant strains), "Trueperella", and some "Enterococcus". It is one of the few antibiotics that works against multidrug resistant "Enterococcus faecalis" and "E. faecium". Activity against gram-negative bacteria includes "Neisseria meningitidis", some "Haemophilus influenzae", and some of the Enterobacteriaceae (though most Enterobacteriaceae and "Pseudomonas" are resistant). Its spectrum of activity is enhanced by co-administration of sulbactam, a drug that inhibits beta lactamase, an enzyme produced by bacteria to inactivate ampicillin and related antibiotics. It is sometimes used in combination with other antibiotics that have different mechanisms of action, like vancomycin, linezolid, daptomycin, and tigecycline.
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Available forms.
Ampicillin can be administered by mouth, an intramuscular injection (shot) or by intravenous infusion. The oral form, available as capsules or oral suspensions, is not given as an initial treatment for severe infections, but rather as a follow-up to an IM or IV injection. For IV and IM injections, ampicillin is kept as a powder that must be reconstituted.
IV injections must be given slowly, as rapid IV injections can lead to convulsive seizures.
Specific populations.
Ampicillin is one of the most used drugs in pregnancy, and has been found to be generally harmless both by the Food and Drug Administration in the U.S. (which classified it as category B) and the Therapeutic Goods Administration in Australia (which classified it as category A). It is the drug of choice for treating "Listeria monocytogenes" in pregnant women, either alone or combined with an aminoglycoside. Pregnancy increases the clearance of ampicillin by up to 50%, and a higher dose is thus needed to reach therapeutic levels.
Ampicillin crosses the placenta and remains in the amniotic fluid at 50–100% of the concentration in maternal plasma; this can lead to high concentrations of ampicillin in the newborn.
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While lactating mothers secrete some ampicillin into their breast milk, the amount is minimal.
In newborns, ampicillin has a longer half-life and lower plasma protein binding. The clearance by the kidneys is lower, as kidney function has not fully developed.
Contraindications.
Ampicillin is contraindicated in those with a hypersensitivity to penicillins, as they can cause fatal anaphylactic reactions. Hypersensitivity reactions can include frequent skin rashes and hives, exfoliative dermatitis, erythema multiforme, and a temporary decrease in both red and white blood cells.
Ampicillin is not recommended in people with concurrent mononucleosis, as over 40% of patients develop a skin rash.
Side effects.
Ampicillin is comparatively less toxic than other antibiotics, and side effects are more likely in those who are sensitive to penicillins and those with a history of asthma or allergies. In very rare cases, it causes severe side effects such as angioedema, anaphylaxis, and "C. difficile" infection (that can range from mild diarrhea to serious pseudomembranous colitis). Some develop black "furry" tongue. Serious adverse effects also include seizures and serum sickness. The most common side effects, experienced by about 10% of users are diarrhea and rash. Less common side effects can be nausea, vomiting, itching, and blood dyscrasias. The gastrointestinal effects, such as hairy tongue, nausea, vomiting, diarrhea, and colitis, are more common with the oral form of penicillin. Other conditions may develop up several weeks after treatment.
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Overdose.
Ampicillin overdose can cause behavioral changes, confusion, blackouts, and convulsions, as well as neuromuscular hypersensitivity, electrolyte imbalance, and kidney failure.
Interactions.
Ampicillin reacts with probenecid and methotrexate to decrease renal excretion. Large doses of ampicillin can increase the risk of bleeding with concurrent use of warfarin and other oral anticoagulants, possibly by inhibiting platelet aggregation. Ampicillin has been said to make oral contraceptives less effective, but this has been disputed. It can be made less effective by other antibiotic, such as chloramphenicol, erythromycin, cephalosporins, and tetracyclines. For example, tetracyclines inhibit protein synthesis in bacteria, reducing the target against which ampicillin acts. If given at the same time as aminoglycosides, it can bind to it and inactivate it. When administered separately, aminoglycosides and ampicillin can potentiate each other instead.
Ampicillin causes skin rashes more often when given with allopurinol.
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Both the live cholera vaccine and live typhoid vaccine can be made ineffective if given with ampicillin. Ampicillin is normally used to treat cholera and typhoid fever, lowering the immunological response that the body has to mount.
Pharmacology.
Mechanism of action.
Ampicillin is in the penicillin group of beta-lactam antibiotics and is part of the aminopenicillin family. It is roughly equivalent to amoxicillin in terms of activity. Ampicillin is able to penetrate gram-positive and some gram-negative bacteria. It differs from penicillin G, or benzylpenicillin, only by the presence of an amino group. This amino group, present on both ampicillin and amoxicillin, helps these antibiotics pass through the pores of the outer membrane of gram-negative bacteria, such as "Escherichia coli", "Proteus mirabilis", "Salmonella enterica", and "Shigella".
Ampicillin acts as an irreversible inhibitor of the enzyme transpeptidase, which is needed by bacteria to make the cell wall. It inhibits the third and final stage of bacterial cell wall synthesis in binary fission, which ultimately leads to cell lysis; therefore, ampicillin is usually bacteriolytic.
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Pharmacokinetics.
Ampicillin is well-absorbed from the GI tract (though food reduces its absorption), and reaches peak concentrations in one to two hours. The bioavailability is around 62% for parenteral routes. Unlike other penicillins, which usually bind 60–90% to plasma proteins, ampicillin binds to only 15–20%.
Ampicillin is distributed through most tissues, though it is concentrated in the liver and kidneys. It can also be found in the cerebrospinal fluid when the meninges become inflamed (such as, for example, meningitis). Some ampicillin is metabolized by hydrolyzing the beta-lactam ring to penicilloic acid, though most of it is excreted unchanged. In the kidneys, it is filtered out mostly by tubular secretion; some also undergoes glomerular filtration, and the rest is excreted in the feces and bile.
Hetacillin and pivampicillin are ampicillin esters that have been developed to increase bioavailability.
History.
Ampicillin has been used extensively to treat bacterial infections since 1961. Until the introduction of ampicillin by the British company Beecham, penicillin therapies had only been effective against gram-positive organisms such as staphylococci and streptococci. Ampicillin (originally branded as "Penbritin") also demonstrated activity against gram-negative organisms such as "H. influenzae", coliforms, and "Proteus" spp.
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Society and culture.
Economics.
Ampicillin is relatively inexpensive. In the United States, it is available as a generic medication.
Veterinary use.
In veterinary medicine, ampicillin is used in cats, dogs, and farm animals to treat:
Horses are generally not treated with oral ampicillin, as they have low bioavailability of beta-lactams.
The half-life in animals is around that same of that in humans (just over an hour). Oral absorption is less than 50% in cats and dogs, and less than 4% in horses.
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Antimicrobial resistance
Antimicrobial resistance (AMR or AR) occurs when microbes evolve mechanisms that protect them from antimicrobials, which are drugs used to treat infections. This resistance affects all classes of microbes, including bacteria (antibiotic resistance), viruses (antiviral resistance), parasites (antiparasitic resistance), and fungi (antifungal resistance). Together, these adaptations fall under the AMR umbrella, posing significant challenges to healthcare worldwide. Misuse and improper management of antimicrobials are primary drivers of this resistance, though it can also occur naturally through genetic mutations and the spread of resistant genes.
Antibiotic resistance, a significant AMR subset, enables bacteria to survive antibiotic treatment, complicating infection management and treatment options. Resistance arises through spontaneous mutation, horizontal gene transfer, and increased selective pressure from antibiotic overuse, both in medicine and agriculture, which accelerates resistance development.
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The burden of AMR is immense, with nearly 5 million annual deaths associated with resistant infections. Infections from AMR microbes are more challenging to treat and often require costly alternative therapies that may have more severe side effects. Preventive measures, such as using narrow-spectrum antibiotics and improving hygiene practices, aim to reduce the spread of resistance. Microbes resistant to multiple drugs are termed multidrug-resistant (MDR) and are sometimes called superbugs.
The World Health Organization (WHO) claims that AMR is one of the top global public health and development threats, estimating that bacterial AMR was directly responsible for 1.27 million global deaths in 2019 and contributed to 4.95 million deaths. Moreover, the WHO and other international bodies warn that AMR could lead to up to 10 million deaths annually by 2050 unless actions are taken. Global initiatives, such as calls for international AMR treaties, emphasize coordinated efforts to limit misuse, fund research, and provide access to necessary antimicrobials in developing nations. However, the COVID-19 pandemic redirected resources and scientific attention away from AMR, intensifying the challenge.
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Definition.
Antimicrobial resistance means a microorganism's resistance to an antimicrobial drug that was once able to treat an infection by that microorganism. A person cannot become resistant to antibiotics. Resistance is a property of the microbe, not a person or other organism infected by a microbe. All types of microbes can develop drug resistance. Thus, there are antibiotic, antifungal, antiviral and antiparasitic resistance.
Antibiotic resistance is a subset of antimicrobial resistance. This more specific resistance is linked to bacteria and thus broken down into two further subsets, microbiological and clinical. Microbiological resistance is the most common and occurs from genes, mutated or inherited, that allow the bacteria to resist the mechanism to kill the microbe associated with certain antibiotics. Clinical resistance is shown through the failure of many therapeutic techniques where the bacteria that are normally susceptible to a treatment become resistant after surviving the outcome of the treatment. In both cases of acquired resistance, the bacteria can pass the genetic catalyst for resistance through horizontal gene transfer: conjugation, transduction, or transformation. This allows the resistance to spread across the same species of pathogen or even similar bacterial pathogens.
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Overview.
WHO report released April 2014 stated, "this serious threat is no longer a prediction for the future, it is happening right now in every region of the world and has the potential to affect anyone, of any age, in any country. Antibiotic resistance—when bacteria change so antibiotics no longer work in people who need them to treat infections—is now a major threat to public health."
Each year, nearly 5 million deaths are associated with AMR globally. In 2019, global deaths attributable to AMR numbered 1.27 million in 2019. That same year, AMR may have contributed to 5 million deaths and one in five people who died due to AMR were children under five years old.
In 2018, WHO considered antibiotic resistance to be one of the biggest threats to global health, food security and development. Deaths attributable to AMR vary by area:
The European Centre for Disease Prevention and Control calculated that in 2015 there were 671,689 infections in the EU and European Economic Area caused by antibiotic-resistant bacteria, resulting in 33,110 deaths. Most were acquired in healthcare settings. In 2019 there were 133,000 deaths caused by AMR.
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Causes.
AMR is driven largely by the misuse and overuse of antimicrobials. Yet, at the same time, many people around the world do not have access to essential antimicrobials. This leads to microbes either evolving a defense against drugs used to treat them, or certain strains of microbes that have a natural resistance to antimicrobials becoming much more prevalent than the ones that are easily defeated with medication. While antimicrobial resistance does occur naturally over time, the use of antimicrobial agents in a variety of settings both within the healthcare industry and outside of has led to antimicrobial resistance becoming increasingly more prevalent.
Although many microbes develop resistance to antibiotics over time though natural mutation, overprescribing and inappropriate prescription of antibiotics have accelerated the problem. It is possible that as many as 1 in 3 prescriptions written for antibiotics are unnecessary. Every year, approximately 154 million prescriptions for antibiotics are written. Of these, up to 46 million are unnecessary or inappropriate for the condition that the patient has. Microbes may naturally develop resistance through genetic mutations that occur during cell division, and although random mutations are rare, many microbes reproduce frequently and rapidly, increasing the chances of members of the population acquiring a mutation that increases resistance. Many individuals stop taking antibiotics when they begin to feel better. When this occurs, it is possible that the microbes that are less susceptible to treatment still remain in the body. If these microbes are able to continue to reproduce, this can lead to an infection by bacteria that are less susceptible or even resistant to an antibiotic.
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Natural occurrence.
AMR is a naturally occurring process. Antimicrobial resistance can evolve naturally due to continued exposure to antimicrobials. Natural selection means that organisms that are able to adapt to their environment, survive, and continue to produce offspring. As a result, the types of microorganisms that are able to survive over time with continued attack by certain antimicrobial agents will naturally become more prevalent in the environment, and those without this resistance will become obsolete.
Some contemporary antimicrobial resistances have also evolved naturally before the use of antimicrobials of human clinical uses. For instance, methicillin-resistance evolved as a pathogen of hedgehogs, possibly as a co-evolutionary adaptation of the pathogen to hedgehogs that are infected by a dermatophyte that naturally produces antibiotics. Also, many soil fungi and bacteria are natural competitors and the original antibiotic penicillin discovered by Alexander Fleming rapidly lost clinical effectiveness in treating humans and, furthermore, none of the other natural penicillins (F, K, N, X, O, U1 or U6) are currently in clinical use.
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Antimicrobial resistance can be acquired from other microbes through swapping genes in a process termed horizontal gene transfer. This means that once a gene for resistance to an antibiotic appears in a microbial community, it can then spread to other microbes in the community, potentially moving from a non-disease causing microbe to a disease-causing microbe. This process is heavily driven by the natural selection processes that happen during antibiotic use or misuse.
Over time, most of the strains of bacteria and infections present will be the type resistant to the antimicrobial agent being used to treat them, making this agent now ineffective to defeat most microbes. With the increased use of antimicrobial agents, there is a speeding up of this natural process.
Self-medication.
In the vast majority of countries, antibiotics can only be prescribed by a doctor and supplied by a pharmacy. Self-medication by consumers is defined as "the taking of medicines on one's own initiative or on another person's suggestion, who is not a certified medical professional", and it has been identified as one of the primary reasons for the evolution of antimicrobial resistance. Self-medication with antibiotics is an unsuitable way of using them but a common practice in resource-constrained countries. The practice exposes individuals to the risk of bacteria that have developed antimicrobial resistance. Many people resort to this out of necessity, when access to a physician is unavailable, or when patients have a limited amount of time or money to see a doctor. This increased access makes it extremely easy to obtain antimicrobials. An example is India, where in the state of Punjab 73% of the population resorted to treating their minor health issues and chronic illnesses through self-medication.
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Self-medication is higher outside the hospital environment, and this is linked to higher use of antibiotics, with the majority of antibiotics being used in the community rather than hospitals. The prevalence of self-medication in low- and middle-income countries (LMICs) ranges from 8.1% to 93%. Accessibility, affordability, and conditions of health facilities, as well as the health-seeking behavior, are factors that influence self-medication in low- and middle-income countries. Two significant issues with self-medication are the lack of knowledge of the public on, firstly, the dangerous effects of certain antimicrobials (for example ciprofloxacin which can cause tendonitis, tendon rupture and aortic dissection) and, secondly, broad microbial resistance and when to seek medical care if the infection is not clearing. In order to determine the public's knowledge and preconceived notions on antibiotic resistance, a screening of 3,537 articles published in Europe, Asia, and North America was done. Of the 55,225 total people surveyed in the articles, 70% had heard of antibiotic resistance previously, but 88% of those people thought it referred to some type of physical change in the human body.
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Clinical misuse.
Clinical misuse by healthcare professionals is another contributor to increased antimicrobial resistance. Studies done in the US show that the indication for treatment of antibiotics, choice of the agent used, and the duration of therapy was incorrect in up to 50% of the cases studied. In 2010 and 2011 about a third of antibiotic prescriptions in outpatient settings in the United States were not necessary. Another study in an intensive care unit in a major hospital in France has shown that 30% to 60% of prescribed antibiotics were unnecessary. These inappropriate uses of antimicrobial agents promote the evolution of antimicrobial resistance by supporting the bacteria in developing genetic alterations that lead to resistance.
According to research conducted in the US that aimed to evaluate physicians' attitudes and knowledge on antimicrobial resistance in ambulatory settings, only 63% of those surveyed reported antibiotic resistance as a problem in their local practices, while 23% reported the aggressive prescription of antibiotics as necessary to avoid failing to provide adequate care. This demonstrates how a majority of doctors underestimate the impact that their own prescribing habits have on antimicrobial resistance as a whole. It also confirms that some physicians may be overly cautious and prescribe antibiotics for both medical or legal reasons, even when clinical indications for use of these medications are not always confirmed. This can lead to unnecessary antimicrobial use, a pattern which may have worsened during the COVID-19 pandemic.
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Studies have shown that common misconceptions about the effectiveness and necessity of antibiotics to treat common mild illnesses contribute to their overuse.
Important to the conversation of antibiotic use is the veterinary medical system. Veterinary oversight is required by law for all medically important antibiotics. Veterinarians use the Pharmacokinetic/pharmacodynamic model (PK/PD) approach to ensuring that the correct dose of the drug is delivered to the correct place at the correct timing.
Pandemics, disinfectants and healthcare systems.
Increased antibiotic use during the early waves of the COVID-19 pandemic may exacerbate this global health challenge. Moreover, pandemic burdens on some healthcare systems may contribute to antibiotic-resistant infections. On the other hand, "increased hand hygiene, decreased international travel, and decreased elective hospital procedures may have reduced AMR pathogen selection and spread in the short term" during the COVID-19 pandemic. The use of disinfectants such as alcohol-based hand sanitizers, and antiseptic hand wash may also have the potential to increase antimicrobial resistance. Extensive use of disinfectants can lead to mutations that induce antimicrobial resistance.
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A 2024 United Nations High-Level Meeting on AMR has pledged to reduce deaths associated with bacterial AMR by 10% over the next six years. In their first major declaration on the issue since 2016, global leaders also committed to raising $100 million to update and implement AMR action plans. However, the final draft of the declaration omitted an earlier target to reduce antibiotic use in animals by 30% by 2030, due to opposition from meat-producing countries and the farming industry. Critics argue this omission is a major weakness, as livestock accounts for around 73% of global sales of antimicrobial agents, including antibiotics, antivirals, and antiparasitics.
Environmental pollution.
Considering the complex interactions between humans, animals and the environment, it is also important to consider the environmental aspects and contributors to antimicrobial resistance. Although there are still some knowledge gaps in understanding the mechanisms and transmission pathways, environmental pollution is considered a significant contributor to antimicrobial resistance.
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Although there are still some knowledge gaps in understanding the mechanisms and transmission pathways, environmental pollution is considered a significant contributor to antimicrobial resistance. Important contributing factors are through "antibiotic residues", "industrial effluents", " agricultural runoffs", "heavy metals", "biocides and pesticides" and "sewage and wastewater" that create reservoirs for resistant genes and bacteria that facilitates the transfer of human pathogens. Unused or expired antibiotics, if not disposed of properly, can enter water systems and soil. Discharge from pharmaceutical manufacturing and other industrial companies can also introduce antibiotics and other chemicals into the environment. These factors allow for creating selective pressure for resistant bacteria. Antibiotics used in livestock and aquaculture can contaminate soil and water, which promotes resistance in environmental microbes. Heavy metals such as zinc, copper and mercury, and also biocides and pesticides, can co- select for antibiotic resistance, enhancing their speed. Heavy metals such as zinc, copper and mercury, and also biocides and pesticides, can co- select for antibiotic resistance, enhancing their speed. Inadequate treatment of sewage and wastewater allows resistant bacteria and genes to spread through water systems.
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Food production.
Livestock.
The antimicrobial resistance crisis also extends to the food industry, specifically with food producing animals. With an ever-increasing human population, there is constant pressure to intensify productivity in many agricultural sectors, including the production of meat as a source of protein. Antibiotics are fed to livestock to act as growth supplements, and a preventive measure to decrease the likelihood of infections.
Farmers typically use antibiotics in animal feed to improve growth rates and prevent infections. However, this is illogical as antibiotics are used to treat infections and not prevent infections. 80% of antibiotic use in the U.S. is for agricultural purposes and about 70% of these are medically important. Overusing antibiotics gives the bacteria time to adapt leaving higher doses or even stronger antibiotics needed to combat the infection. Though antibiotics for growth promotion were banned throughout the EU in 2006, 40 countries worldwide still use antibiotics to promote growth.
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This can result in the transfer of resistant bacterial strains into the food that humans eat, causing potentially fatal transfer of disease. While the practice of using antibiotics as growth promoters does result in better yields and meat products, it is a major issue and needs to be decreased in order to prevent antimicrobial resistance. Though the evidence linking antimicrobial usage in livestock to antimicrobial resistance is limited, the World Health Organization Advisory Group on Integrated Surveillance of Antimicrobial Resistance strongly recommended the reduction of use of medically important antimicrobials in livestock. Additionally, the Advisory Group stated that such antimicrobials should be expressly prohibited for both growth promotion and disease prevention in food producing animals.
By mapping antimicrobial consumption in livestock globally, it was predicted that in 228 countries there would be a total 67% increase in consumption of antibiotics by livestock by 2030. In some countries such as Brazil, Russia, India, China, and South Africa it is predicted that a 99% increase will occur. Several countries have restricted the use of antibiotics in livestock, including Canada, China, Japan, and the US. These restrictions are sometimes associated with a reduction of the prevalence of antimicrobial resistance in humans.
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In the United States the Veterinary Feed Directive went into practice in 2017 dictating that "All medically important antibiotics to be used in feed or water for food animal species require a veterinary feed directive (VFD) or a prescription."
Pesticides.
Most pesticides protect crops against insects and plants, but in some cases antimicrobial pesticides are used to protect against various microorganisms such as bacteria, viruses, fungi, algae, and protozoa. The overuse of many pesticides in an effort to have a higher yield of crops has resulted in many of these microbes evolving a tolerance against these antimicrobial agents. Currently there are over 4000 antimicrobial pesticides registered with the US Environmental Protection Agency (EPA) and sold to market, showing the widespread use of these agents. It is estimated that for every single meal a person consumes, 0.3 g of pesticides is used, as 90% of all pesticide use is in agriculture. A majority of these products are used to help defend against the spread of infectious diseases, and hopefully protect public health. But out of the large amount of pesticides used, it is also estimated that less than 0.1% of those antimicrobial agents, actually reach their targets. That leaves over 99% of all pesticides used available to contaminate other resources. In soil, air, and water these antimicrobial agents are able to spread, coming in contact with more microorganisms and leading to these microbes evolving mechanisms to tolerate and further resist pesticides. The use of antifungal azole pesticides that drive environmental azole resistance have been linked to azole resistance cases in the clinical setting. The same issues confront the novel antifungal classes (e.g. orotomides) which are again being used in both the clinic and agriculture.
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Wild birds.
Wildlife, including wild and migratory birds, serve as a reservoir for zoonotic disease and antimicrobial-resistant organisms. Birds are a key link between the transmission of zoonotic diseases to human populations. By the same token, increased contact between wild birds and human populations (including domesticated animals), has increased the amount of anti-microbial resistance (AMR) to the bird population. The introduction of AMR to wild birds positively correlates with human pollution and increased human contact. Additionally, wild birds can participate in horizontal gene transfer with bacteria, leading to the transmission of antibiotic-resistant genes (ARG).
For simplicity, wild bird populations can be divided into two major categories, wild sedentary birds and wild migrating birds. Wild sedentary bird exposure to AMR is through increased contact with densely populated areas, human waste, domestic animals, and domestic animal/livestock waste. Wild migrating birds interact with sedentary birds in different environments along their migration route. This increases the rate and diversity of AMR across varying ecosystems.
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Neglect of wildlife in the global discussions surrounding health security and AMR, creates large barriers to true AMR surveillance. The surveillance of anti-microbial resistant organisms in wild birds is a potential metric for the rate of AMR in the environment. This surveillance also allows for further investigation into the transmission routs between different ecosystems and human populations (including domesticated animals and livestock). Such information gathered from wild bird biomes, can help identify patterns of diseased transmission and better target interventions. These targeted interventions can inform the use of antimicrobial agents and reduce the persistence of multi-drug resistant organisms.
Gene transfer from ancient microorganisms.
Permafrost is a term used to refer to any ground that remained frozen for two years or more, with the oldest known examples continuously frozen for around 700,000 years. In the recent decades, permafrost has been rapidly thawing due to climate change. The cold preserves any organic matter inside the permafrost, and it is possible for microorganisms to resume their life functions once it thaws. While some common pathogens such as influenza, smallpox or the bacteria associated with pneumonia have failed to survive intentional attempts to revive them, more cold-adapted microorganisms such as anthrax, or several ancient plant and amoeba viruses, have successfully survived prolonged thaw.
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Some scientists have argued that the inability of known causative agents of contagious diseases to survive being frozen and thawed makes this threat unlikely. Instead, there have been suggestions that when modern pathogenic bacteria interact with the ancient ones, they may, through horizontal gene transfer, pick up genetic sequences which are associated with antimicrobial resistance, exacerbating an already difficult issue. Antibiotics to which permafrost bacteria have displayed at least some resistance include chloramphenicol, streptomycin, kanamycin, gentamicin, tetracycline, spectinomycin and neomycin. However, other studies show that resistance levels in ancient bacteria to modern antibiotics remain lower than in the contemporary bacteria from the active layer of thawed ground above them, which may mean that this risk is "no greater" than from any other soil.
Prevention.
There have been increasing public calls for global collective action to address the threat, including a proposal for an international treaty on antimicrobial resistance. Further detail and attention is still needed in order to recognize and measure trends in resistance on the international level; the idea of a global tracking system has been suggested but implementation has yet to occur. A system of this nature would provide insight to areas of high resistance as well as information necessary for evaluating programs, introducing interventions and other changes made to fight or reverse antibiotic resistance.
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Duration of antimicrobials.
Delaying or minimizing the use of antibiotics for certain conditions may help safely reduce their use. Antimicrobial treatment duration should be based on the infection and other health problems a person may have. For many infections once a person has improved there is little evidence that stopping treatment causes more resistance. Some, therefore, feel that stopping early may be reasonable in some cases. Other infections, however, do require long courses regardless of whether a person feels better.
Delaying antibiotics for ailments such as a sore throat and otitis media may have no difference in the rate of complications compared with immediate antibiotics, for example. When treating respiratory tract infections, clinical judgement is required as to the appropriate treatment (delayed or immediate antibiotic use).
The study, "Shorter and Longer Antibiotic Durations for Respiratory Infections: To Fight Antimicrobial Resistance—A Retrospective Cross-Sectional Study in a Secondary Care Setting in the UK," highlights the urgency of reevaluating antibiotic treatment durations amidst the global challenge of antimicrobial resistance (AMR). It investigates the effectiveness of shorter versus longer antibiotic regimens for respiratory tract infections (RTIs) in a UK secondary care setting, emphasizing the need for evidence-based prescribing practices to optimize patient outcomes and combat AMR.
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Monitoring and mapping.
There are multiple national and international monitoring programs for drug-resistant threats, including methicillin-resistant "Staphylococcus aureus" (MRSA), vancomycin-resistant "S. aureus" (VRSA), extended spectrum beta-lactamase (ESBL) producing Enterobacterales, vancomycin-resistant "Enterococcus" (VRE), and multidrug-resistant "Acinetobacter baumannii" (MRAB).
ResistanceOpen is an online global map of antimicrobial resistance developed by HealthMap which displays aggregated data on antimicrobial resistance from publicly available and user submitted data. The website can display data for a radius from a location. Users may submit data from antibiograms for individual hospitals or laboratories. European data is from the EARS-Net (European Antimicrobial Resistance Surveillance Network), part of the ECDC. ResistanceMap is a website by the Center for Disease Dynamics, Economics & Policy and provides data on antimicrobial resistance on a global level.
The WHO's AMR global action plan also recommends antimicrobial resistance surveillance in animals. Initial steps in the EU for establishing the veterinary counterpart EARS-Vet (EARS-Net for veterinary medicine) have been made. AMR data from pets in particular is scarce, but needed to support antibiotic stewardship in veterinary medicine.
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By comparison there is a lack of national and international monitoring programs for antifungal resistance.
Limiting antimicrobial use in humans.
Antimicrobial stewardship programmes appear useful in reducing rates of antimicrobial resistance. The antimicrobial stewardship program will also provide pharmacists with the knowledge to educate patients that antibiotics will not work for a virus for example.
Excessive antimicrobial use has become one of the top contributors to the evolution of antimicrobial resistance. Since the beginning of the antimicrobial era, antimicrobials have been used to treat a wide range of infectious diseases. Overuse of antimicrobials has become the primary cause of rising levels of antimicrobial resistance. The main problem is that doctors are willing to prescribe antimicrobials to ill-informed individuals who believe that antimicrobials can cure nearly all illnesses, including viral infections like the common cold. In an analysis of drug prescriptions, 36% of individuals with a cold or an upper respiratory infection (both usually viral in origin) were given prescriptions for antibiotics. These prescriptions accomplished nothing other than increasing the risk of further evolution of antibiotic resistant bacteria. Using antimicrobials without prescription is another driving force leading to the overuse of antibiotics to self-treat diseases like the common cold, cough, fever, and dysentery resulting in an epidemic of antibiotic resistance in countries like Bangladesh, risking its spread around the globe. Introducing strict antibiotic stewardship in the outpatient setting to reduce inappropriate prescribing of antibiotics may reduce the emerging bacterial resistance.
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The WHO AWaRe (Access, Watch, Reserve) guidance and antibiotic book has been introduced to guide antibiotic choice for the 30 most common infections in adults and children to reduce inappropriate prescribing in primary care and hospitals. Narrow-spectrum antibiotics are preferred due to their lower resistance potential, and broad-spectrum antibiotics are only recommended for people with more severe symptoms. Some antibiotics are more likely to confer resistance, so are kept as reserve antibiotics in the AWaRe book.
Various diagnostic strategies have been employed to prevent the overuse of antifungal therapy in the clinic, proving a safe alternative to empirical antifungal therapy, and thus underpinning antifungal stewardship schemes.
At the hospital level.
Antimicrobial stewardship teams in hospitals are encouraging optimal use of antimicrobials. The goals of antimicrobial stewardship are to help practitioners pick the right drug at the right dose and duration of therapy while preventing misuse and minimizing the development of resistance. Stewardship interventions may reduce the length of stay by an average of slightly over 1 day while not increasing the risk of death. Dispensing, to discharged in-house patients, the exact number of antibiotic pharmaceutical units necessary to complete an ongoing treatment can reduce antibiotic leftovers within the community as community pharmacies can have antibiotic package inefficiencies.
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At the primary care level.
Given the volume of care provided in primary care (general practice), recent strategies have focused on reducing unnecessary antimicrobial prescribing in this setting. Simple interventions, such as written information explaining when taking antibiotics is not necessary, for example in common infections of the upper respiratory tract, have been shown to reduce antibiotic prescribing. Various tools are also available to help professionals decide if prescribing antimicrobials is necessary.
Parental expectations, driven by the worry for their children's health, can influence how often children are prescribed antibiotics. Parents often rely on their clinician for advice and reassurance. However a lack of plain language information and not having adequate time for consultation negatively impacts this relationship. In effect parents often rely on past experiences in their expectations rather than reassurance from the clinician. Adequate time for consultation and plain language information can help parents make informed decisions and avoid unnecessary antibiotic use.
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Parents play a critical role in reducing unnecessary antibiotic use, particularly during cold and flu season when children frequently experience respiratory illnesses. Many of these illnesses are caused by viruses, such as colds or the flu, which antibiotics cannot treat. Misusing antibiotics in these situations not only fails to benefit the child but also contributes to the emergence of antibiotic-resistant bacteria, posing a broader public health threat. To address parental concerns and reduce inappropriate prescribing, healthcare providers can offer plain-language explanations about the difference between bacterial and viral infections, alongside clear guidance on managing viral illnesses without antibiotics. Vaccinations also play a vital role in reducing the incidence of serious bacterial infections that may require antibiotic treatment, thereby helping to preserve the effectiveness of existing antibiotics. Schools further amplify the spread of infections due to close contact and shared surfaces, underscoring the importance of hygiene practices like regular handwashing, covering coughs, and staying home when unwell. These preventive measures not only reduce the need for antibiotics but also lower the overall risk of resistant bacteria spreading within communities.
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The prescriber should closely adhere to the five rights of drug administration: the right patient, the right drug, the right dose, the right route, and the right time. Microbiological samples should be taken for culture and sensitivity testing before treatment when indicated and treatment potentially changed based on the susceptibility report.
Health workers and pharmacists can help tackle antibiotic resistance by: enhancing infection prevention and control; only prescribing and dispensing antibiotics when they are truly needed; prescribing and dispensing the right antibiotic(s) to treat the illness. A unit dose system implemented in community pharmacies can also reduce antibiotic leftovers at households.
At the individual level.
People can help tackle resistance by using antibiotics only when infected with a bacterial infection and prescribed by a doctor; completing the full prescription even if the user is feeling better, never sharing antibiotics with others, or using leftover prescriptions. Taking antibiotics when not needed won't help the user, but instead give bacteria the option to adapt and leave the user with the side effects that come with the certain type of antibiotic. The CDC recommends that you follow these behaviors so that you avoid these negative side effects and keep the community safe from spreading drug-resistant bacteria. Practicing basic bacterial infection prevention courses, such as hygiene, also helps to prevent the spread of antibiotic-resistant bacteria.
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Water, sanitation, hygiene.
Infectious disease control through improved water, sanitation and hygiene (WASH) infrastructure needs to be included in the antimicrobial resistance (AMR) agenda. The "Interagency Coordination Group on Antimicrobial Resistance" stated in 2018 that "the spread of pathogens through unsafe water results in a high burden of gastrointestinal disease, increasing even further the need for antibiotic treatment." This is particularly a problem in developing countries where the spread of infectious diseases caused by inadequate WASH standards is a major driver of antibiotic demand. Growing usage of antibiotics together with persistent infectious disease levels have led to a dangerous cycle in which reliance on antimicrobials increases while the efficacy of drugs diminishes. The proper use of infrastructure for water, sanitation and hygiene (WASH) can result in a 47–72 percent decrease of diarrhea cases treated with antibiotics depending on the type of intervention and its effectiveness. A reduction of the diarrhea disease burden through improved infrastructure would result in large decreases in the number of diarrhea cases treated with antibiotics. This was estimated as ranging from 5 million in Brazil to up to 590 million in India by the year 2030. The strong link between increased consumption and resistance indicates that this will directly mitigate the accelerating spread of AMR. Sanitation and water for all by 2030 is Goal Number 6 of the Sustainable Development Goals.
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An increase in hand washing compliance by hospital staff results in decreased rates of resistant organisms.
Water supply and sanitation infrastructure in health facilities offer significant co-benefits for combatting AMR, and investment should be increased. There is much room for improvement: WHO and UNICEF estimated in 2015 that globally 38% of health facilities did not have a source of water, nearly 19% had no toilets and 35% had no water and soap or alcohol-based hand rub for handwashing.
Industrial wastewater treatment.
Manufacturers of antimicrobials need to improve the treatment of their wastewater (by using industrial wastewater treatment processes) to reduce the release of residues into the environment.
Limiting antimicrobial use in animals and farming.
It is established that the use of antibiotics in animal husbandry can give rise to AMR resistances in bacteria found in food animals to the antibiotics being administered (through injections or medicated feeds). For this reason only antimicrobials that are deemed "not-clinically relevant" are used in these practices.
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Unlike resistance to antibacterials, antifungal resistance can be driven by arable farming, currently there is no regulation on the use of similar antifungal classes in agriculture and the clinic.
Recent studies have shown that the prophylactic use of "non-priority" or "non-clinically relevant" antimicrobials in feeds can potentially, under certain conditions, lead to co-selection of environmental AMR bacteria with resistance to medically important antibiotics. The possibility for co-selection of AMR resistances in the food chain pipeline may have far-reaching implications for human health.
Country examples.
Europe.
In 1997, European Union health ministers voted to ban avoparcin and four additional antibiotics used to promote animal growth in 1999. In 2006 a ban on the use of antibiotics in European feed, with the exception of two antibiotics in poultry feeds, became effective. In Scandinavia, there is evidence that the ban has led to a lower prevalence of antibiotic resistance in (nonhazardous) animal bacterial populations. As of 2004, several European countries established a decline of antimicrobial resistance in humans through limiting the use of antimicrobials in agriculture and food industries without jeopardizing animal health or economic cost.
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United States.
The United States Department of Agriculture (USDA) and the Food and Drug Administration (FDA) collect data on antibiotic use in humans and in a more limited fashion in animals. About 80% of antibiotic use in the U.S. is for agriculture purposes, and about 70% of these are medically important. This gives reason for concern about the antibiotic resistance crisis in the U.S. and more reason to monitor it. The FDA first determined in 1977 that there is evidence of emergence of antibiotic-resistant bacterial strains in livestock. The long-established practice of permitting OTC sales of antibiotics (including penicillin and other drugs) to lay animal owners for administration to their own animals nonetheless continued in all states.
In 2000, the FDA announced their intention to revoke approval of fluoroquinolone use in poultry production because of substantial evidence linking it to the emergence of fluoroquinolone-resistant "Campylobacter" infections in humans. Legal challenges from the food animal and pharmaceutical industries delayed the final decision to do so until 2006. Fluroquinolones have been banned from extra-label use in food animals in the USA since 2007. However, they remain widely used in companion and exotic animals.
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Global action plans and awareness.
At the 79th United Nations General Assembly High-Level Meeting on AMR on 26 September 2024, world leaders approved a political declaration committing to a clear set of targets and actions, including reducing the estimated 4.95 million human deaths associated with bacterial AMR annually by 10% by 2030.
The increasing interconnectedness of the world and the fact that new classes of antibiotics have not been developed and approved for more than 25 years highlight the extent to which antimicrobial resistance is a global health challenge. A global action plan to tackle the growing problem of resistance to antibiotics and other antimicrobial medicines was endorsed at the Sixty-eighth World Health Assembly in May 2015. One of the key objectives of the plan is to improve awareness and understanding of antimicrobial resistance through effective communication, education and training. This global action plan developed by the World Health Organization was created to combat the issue of antimicrobial resistance and was guided by the advice of countries and key stakeholders. The WHO's global action plan is composed of five key objectives that can be targeted through different means, and represents countries coming together to solve a major problem that can have future health consequences. These objectives are as follows:
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Steps towards progress
Antibiotic Awareness Week.
The World Health Organization has promoted the first World Antibiotic Awareness Week running from 16 to 22 November 2015. The aim of the week is to increase global awareness of antibiotic resistance. It also wants to promote the correct usage of antibiotics across all fields in order to prevent further instances of antibiotic resistance.
World Antibiotic Awareness Week has been held every November since 2015. For 2017, the Food and Agriculture Organization of the United Nations (FAO), the World Health Organization (WHO) and the World Organisation for Animal Health (OIE) are together calling for responsible use of antibiotics in humans and animals to reduce the emergence of antibiotic resistance.
United Nations
In 2016 the Secretary-General of the United Nations convened the Interagency Coordination Group (IACG) on Antimicrobial Resistance. The IACG worked with international organizations and experts in human, animal, and plant health to create a plan to fight antimicrobial resistance. Their report released in April 2019 highlights the seriousness of antimicrobial resistance and the threat it poses to world health. It suggests five recommendations for member states to follow in order to tackle this increasing threat. The IACG recommendations are as follows:
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One Health Approach.
The One Health approach recognizes that human, animal, and environmental health are interconnected in the development and spread of antimicrobial resistance (AMR). Key strategies include:
Mechanisms and organisms.
Bacteria.
The five main mechanisms by which bacteria exhibit resistance to antibiotics are:
There are several different types of germs that have developed a resistance over time.
The six pathogens causing most deaths associated with resistance are "Escherichia coli", "Staphylococcus aureus, Klebsiella pneumoniae, Streptococcus pneumoniae, Acinetobacter baumannii", and "Pseudomonas aeruginosa". They were responsible for 929,000 deaths attributable to resistance and 3.57 million deaths associated with resistance in 2019.
Penicillinase-producing "Neisseria gonorrhoeae" developed a resistance to penicillin in 1976. Another example is Azithromycin-resistant "Neisseria gonorrhoeae", which developed a resistance to azithromycin in 2011.
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.
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Some bacteria are naturally resistant to certain antibiotics; for example, gram-negative bacteria are resistant to most β-lactam antibiotics due to the presence of β-lactamase. Antibiotic resistance can also be acquired as a result of either genetic mutation or horizontal gene transfer. Although mutations are rare, with spontaneous mutations in the pathogen genome occurring at a rate of about 1 in 105 to 1 in 108 per chromosomal replication, the fact that bacteria reproduce at a high rate allows for the effect to be significant. Given that lifespans and production of new generations can be on a timescale of mere hours, a new (de novo) mutation in a parent cell can quickly become an inherited mutation of widespread prevalence, resulting in the microevolution of a fully resistant colony. However, chromosomal mutations also confer a cost of fitness. For example, a ribosomal mutation may protect a bacterial cell by changing the binding site of an antibiotic but may result in slower growth rate. Moreover, some adaptive mutations can propagate not only through inheritance but also through horizontal gene transfer. The most common mechanism of horizontal gene transfer is the transferring of plasmids carrying antibiotic resistance genes between bacteria of the same or different species via conjugation. However, bacteria can also acquire resistance through transformation, as in "Streptococcus pneumoniae" uptaking of naked fragments of extracellular DNA that contain antibiotic resistance genes to streptomycin, through transduction, as in the bacteriophage-mediated transfer of tetracycline resistance genes between strains of "S. pyogenes", or through gene transfer agents, which are particles produced by the host cell that resemble bacteriophage structures and are capable of transferring DNA.
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Antibiotic resistance can be introduced artificially into a microorganism through laboratory protocols, sometimes used as a selectable marker to examine the mechanisms of gene transfer or to identify individuals that absorbed a piece of DNA that included the resistance gene and another gene of interest.
Recent findings show no necessity of large populations of bacteria for the appearance of antibiotic resistance. Small populations of "Escherichia coli" in an antibiotic gradient can become resistant. Any heterogeneous environment with respect to nutrient and antibiotic gradients may facilitate antibiotic resistance in small bacterial populations. Researchers hypothesize that the mechanism of resistance evolution is based on four SNP mutations in the genome of "E. coli" produced by the gradient of antibiotic.
In one study, which has implications for space microbiology, a non-pathogenic strain "E. coli" MG1655 was exposed to trace levels of the broad spectrum antibiotic chloramphenicol, under simulated microgravity (LSMMG, or Low Shear Modeled Microgravity) over 1000 generations. The adapted strain acquired resistance to not only chloramphenicol, but also cross-resistance to other antibiotics; this was in contrast to the observation on the same strain, which was adapted to over 1000 generations under LSMMG, but without any antibiotic exposure; the strain in this case did not acquire any such resistance. Thus, irrespective of where they are used, the use of an antibiotic would likely result in persistent resistance to that antibiotic, as well as cross-resistance to other antimicrobials.
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In recent years, the emergence and spread of β-lactamases called carbapenemases has become a major health crisis. One such carbapenemase is New Delhi metallo-beta-lactamase 1 (NDM-1), an enzyme that makes bacteria resistant to a broad range of beta-lactam antibiotics. The most common bacteria that make this enzyme are gram-negative such as "E. coli" and "Klebsiella pneumoniae", but the gene for NDM-1 can spread from one strain of bacteria to another by horizontal gene transfer.
Viruses.
Specific antiviral drugs are used to treat some viral infections. These drugs prevent viruses from reproducing by inhibiting essential stages of the virus's replication cycle in infected cells. Antivirals are used to treat HIV, hepatitis B, hepatitis C, influenza, herpes viruses including varicella zoster virus, cytomegalovirus and Epstein–Barr virus. With each virus, some strains have become resistant to the administered drugs.
Antiviral drugs typically target key components of viral reproduction; for example, oseltamivir targets influenza neuraminidase, while guanosine analogs inhibit viral DNA polymerase. Resistance to antivirals is thus acquired through mutations in the genes that encode the protein targets of the drugs.
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Resistance to HIV antivirals is problematic, and even multi-drug resistant strains have evolved. One source of resistance is that many current HIV drugs, including NRTIs and NNRTIs, target reverse transcriptase; however, HIV-1 reverse transcriptase is highly error prone and thus mutations conferring resistance arise rapidly. Resistant strains of the HIV virus emerge rapidly if only one antiviral drug is used. Using three or more drugs together, termed combination therapy, has helped to control this problem, but new drugs are needed because of the continuing emergence of drug-resistant HIV strains.
Fungi.
Infections by fungi are a cause of high morbidity and mortality in immunocompromised persons, such as those with HIV/AIDS, tuberculosis or receiving chemotherapy. The fungi "Candida", "Cryptococcus neoformans" and "Aspergillus fumigatus" cause most of these infections and antifungal resistance occurs in all of them. Multidrug resistance in fungi is increasing because of the widespread use of antifungal drugs to treat infections in immunocompromised individuals and the use of some agricultural antifungals. Antifungal resistant disease is associated with increased mortality.
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Some fungi (e.g. Candida krusei and fluconazole) exhibit intrinsic resistance to certain antifungal drugs or classes, whereas some species develop antifungal resistance to external pressures. Antifungal resistance is a One Health concern, driven by multiple extrinsic factors, including extensive fungicidal use, overuse of clinical antifungals, environmental change and host factors.
In the USA fluconazole-resistant Candida species and azole resistance in Aspergillus fumigatus have been highlighted as a growing threat.
More than 20 species of "Candida" can cause candidiasis infection, the most common of which is "Candida albicans". "Candida" yeasts normally inhabit the skin and mucous membranes without causing infection. However, overgrowth of "Candida" can lead to candidiasis. Some "Candida" species (e.g. "Candida glabrata)" are becoming resistant to first-line and second-line antifungal agents such as echinocandins and azoles.
The emergence of "Candida auris" as a potential human pathogen that sometimes exhibits multi-class antifungal drug resistance is concerning and has been associated with several outbreaks globally. The WHO has released a priority fungal pathogen list, including pathogens with antifungal resistance.
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The identification of antifungal resistance is undermined by limited classical diagnosis of infection, where a culture is lacking, preventing susceptibility testing. National and international surveillance schemes for fungal disease and antifungal resistance are limited, hampering the understanding of the disease burden and associated resistance. The application of molecular testing to identify genetic markers associating with resistance may improve the identification of antifungal resistance, but the diversity of mutations associated with resistance is increasing across the fungal species causing infection. In addition, a number of resistance mechanisms depend on up-regulation of selected genes (for instance reflux pumps) rather than defined mutations that are amenable to molecular detection.
Due to the limited number of antifungals in clinical use and the increasing global incidence of antifungal resistance, using the existing antifungals in combination might be beneficial in some cases but further research is needed. Similarly, other approaches that might help to combat the emergence of antifungal resistance could rely on the development of host-directed therapies such as immunotherapy or vaccines.
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Parasites.
The protozoan parasites that cause the diseases malaria, trypanosomiasis, toxoplasmosis, cryptosporidiosis and leishmaniasis are important human pathogens.
Malarial parasites that are resistant to the drugs that are currently available to infections are common and this has led to increased efforts to develop new drugs. Resistance to recently developed drugs such as artemisinin has also been reported. The problem of drug resistance in malaria has driven efforts to develop vaccines.
Trypanosomes are parasitic protozoa that cause African trypanosomiasis and Chagas disease (American trypanosomiasis). There are no vaccines to prevent these infections so drugs such as pentamidine and suramin, benznidazole and nifurtimox are used to treat infections. These drugs are effective but infections caused by resistant parasites have been reported.
Leishmaniasis is caused by protozoa and is an important public health problem worldwide, especially in sub-tropical and tropical countries. Drug resistance has "become a major concern".
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Global and genomic data.
In 2022, genomic epidemiologists reported results from a global survey of antimicrobial resistance via genomic wastewater-based epidemiology, finding large regional variations, providing maps, and suggesting resistance genes are also passed on between microbial species that are not closely related.<ref name="10.1038/s41467-022-34312-7"></ref> The WHO provides the Global Antimicrobial Resistance and Use Surveillance System (GLASS) reports which summarize annual (e.g. 2020's) data on international AMR, also including an interactive dashboard.
Epidemiology.
United Kingdom.
Public Health England reported that the total number of antibiotic resistant infections in England rose by 9% from 55,812 in 2017 to 60,788 in 2018, but antibiotic consumption had fallen by 9% from 20.0 to 18.2 defined daily doses per 1,000 inhabitants per day between 2014 and 2018.
United States.
The Centers for Disease Control and Prevention reported that more than 2.8 million cases of antibiotic resistance have been reported. However, in 2019 overall deaths from antibiotic-resistant infections decreased by 18% and deaths in hospitals decreased by 30%.
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The COVID pandemic caused a reversal of much of the progress made on attenuating the effects of antibiotic resistance, resulting in more antibiotic use, more resistant infections, and less data on preventive action. Hospital-onset infections and deaths both increased by 15% in 2020, and significantly higher rates of infections were reported for 4 out of 6 types of healthcare associated infections.
History.
The 1950s to 1970s represented the golden age of antibiotic discovery, where countless new classes of antibiotics were discovered to treat previously incurable diseases such as tuberculosis and syphilis. However, since that time the discovery of new classes of antibiotics has been almost nonexistent, and represents a situation that is especially problematic considering the resiliency of bacteria shown over time and the continued misuse and overuse of antibiotics in treatment.
Already in 1940, in their letter to the editor of Nature journal, Abraham and Chain identified the enzyme penicillinase as responsible for the deactivation of penicillin in penicillin-resistant bacteria. This discovery was the first step in understanding the mechanisms of microbial resistance to β-lactam antibiotics. The phenomenon of antimicrobial resistance caused by overuse of antibiotics was predicted as early as 1945 by Alexander Fleming who said "The time may come when penicillin can be bought by anyone in the shops. Then there is the danger that the ignorant man may easily under-dose himself and by exposing his microbes to nonlethal quantities of the drug make them resistant." Without the creation of new and stronger antibiotics an era where common infections and minor injuries can kill, and where complex procedures such as surgery and chemotherapy become too risky, is a very real possibility. Antimicrobial resistance can lead to epidemics of enormous proportions if preventive actions are not taken. In this day and age current antimicrobial resistance leads to longer hospital stays, higher medical costs, and increased mortality.
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Society and culture.
Innovation policy.
Since the mid-1980s pharmaceutical companies have invested in medications for cancer or chronic disease that have greater potential to make money and have "de-emphasized or dropped development of antibiotics". On 20 January 2016 at the World Economic Forum in Davos, Switzerland, more than "80 pharmaceutical and diagnostic companies" from around the world called for "transformational commercial models" at a global level to spur research and development on antibiotics and on the "enhanced use of diagnostic tests that can rapidly identify the infecting organism". A number of countries are considering or implementing delinked payment models for new antimicrobials whereby payment is based on value rather than volume of drug sales. This offers the opportunity to pay for valuable new drugs even if they are reserved for use in relatively rare drug resistant infections.
Legal frameworks.
Some global health scholars have argued that a global, legal framework is needed to prevent and control antimicrobial resistance. For instance, binding global policies could be used to create antimicrobial use standards, regulate antibiotic marketing, and strengthen global surveillance systems. Ensuring compliance of involved parties is a challenge. Global antimicrobial resistance policies could take lessons from the environmental sector by adopting strategies that have made international environmental agreements successful in the past such as: sanctions for non-compliance, assistance for implementation, majority vote decision-making rules, an independent scientific panel, and specific commitments.
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United States.
For the United States 2016 budget, U.S. president Barack Obama proposed to nearly double the amount of federal funding to "combat and prevent" antibiotic resistance to more than $1.2 billion. Many international funding agencies like USAID, DFID, SIDA and Gates Foundation have pledged money for developing strategies to counter antimicrobial resistance.
On 27 March 2015, the White House released a comprehensive plan to address the increasing need for agencies to combat the rise of antibiotic-resistant bacteria. The Task Force for Combating Antibiotic-Resistant Bacteria developed "The National Action Plan for Combating Antibiotic-Resistant Bacteria" with the intent of providing a roadmap to guide the US in the antibiotic resistance challenge and with hopes of saving many lives. This plan outlines steps taken by the Federal government over the next five years needed in order to prevent and contain outbreaks of antibiotic-resistant infections; maintain the efficacy of antibiotics already on the market; and to help to develop future diagnostics, antibiotics, and vaccines.
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The Action Plan was developed around five goals with focuses on strengthening health care, public health veterinary medicine, agriculture, food safety and research, and manufacturing. These goals, as listed by the White House, are as follows:
The following are goals set to meet by 2020:
Current Status of AMR in the U.S.
As of 2023, antimicrobial resistance (AMR) remains a significant public health threat in the United States. According to the Centers for Disease Control and Prevention's 2023 Report on Antibiotic Resistance Threats, over 2.8 million antibiotic-resistant infections occur in the U.S. each year, leading to at least 35,000 deaths annually. Among the most concerning resistant pathogens are Carbapenem-resistant Enterobacteriaceae (CRE), Methicillin-resistant Staphylococcus aureus (MRSA), and Clostridioides difficile (C. diff), all of which continue to be responsible for severe healthcare-associated infections (HAIs).
The COVID-19 pandemic led to a significant disruption in healthcare, with an increase in the use of antibiotics during the treatment of viral infections. This rise in antibiotic prescribing, coupled with overwhelmed healthcare systems, contributed to a resurgence in AMR during the pandemic years. A 2021 CDC report identified a sharp increase in HAIs caused by resistant pathogens in COVID-19 patients, a trend that has persisted into 2023. Recent data suggest that although antibiotic use has decreased since the pandemic, some resistant pathogens remain prevalent in healthcare settings.
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The CDC has also expanded its Get Ahead of Sepsis campaign in 2023, focusing on raising awareness of AMR's role in sepsis and promoting the judicious use of antibiotics in both healthcare and community settings. This initiative has reached millions through social media, healthcare facilities, and public health outreach, aiming to educate the public on the importance of preventing infections and reducing antibiotic misuse.
Policies.
According to World Health Organization, policymakers can help tackle resistance by strengthening resistance-tracking and laboratory capacity and by regulating and promoting the appropriate use of medicines. Policymakers and industry can help tackle resistance by: fostering innovation and research and development of new tools; and promoting cooperation and information sharing among all stakeholders.
The U.S. government continues to prioritize AMR mitigation through policy and legislation. In 2023, the National Action Plan for Combating Antibiotic-Resistant Bacteria (CARB) 2023-2028 was released, outlining strategic objectives for reducing antibiotic-resistant infections, advancing infection prevention, and accelerating research on new antibiotics. The plan also emphasizes the importance of improving antibiotic stewardship across healthcare, agriculture, and veterinary settings.
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Furthermore, the PASTEUR Act (Pioneering Antimicrobial Subscriptions to End Upsurging Resistance) has gained momentum in Congress. If passed, the bill would create a subscription-based payment model to incentivize the development of new antimicrobial drugs, while supporting antimicrobial stewardship programs to reduce the misuse of existing antibiotics. This legislation is considered a critical step toward addressing the economic barriers to developing new antimicrobials.
Policy evaluation.
Measuring the costs and benefits of strategies to combat AMR is difficult and policies may only have effects in the distant future. In other infectious diseases this problem has been addressed by using mathematical models. More research is needed to understand how AMR develops and spreads so that mathematical modelling can be used to anticipate the likely effects of different policies.
Further research.
Rapid testing and diagnostics.
Distinguishing infections requiring antibiotics from self-limiting ones is clinically challenging. In order to guide appropriate use of antibiotics and prevent the evolution and spread of antimicrobial resistance, diagnostic tests that provide clinicians with timely, actionable results are needed.
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Acute febrile illness is a common reason for seeking medical care worldwide and a major cause of morbidity and mortality. In areas with decreasing malaria incidence, many febrile patients are inappropriately treated for malaria, and in the absence of a simple diagnostic test to identify alternative causes of fever, clinicians presume that a non-malarial febrile illness is most likely a bacterial infection, leading to inappropriate use of antibiotics. Multiple studies have shown that the use of malaria rapid diagnostic tests without reliable tools to distinguish other fever causes has resulted in increased antibiotic use.
Antimicrobial susceptibility testing (AST) can facilitate a precision medicine approach to treatment by helping clinicians to prescribe more effective and targeted antimicrobial therapy. At the same time with traditional phenotypic AST it can take 12 to 48 hours to obtain a result due to the time taken for organisms to grow on/in culture media. Rapid testing, possible from molecular diagnostics innovations, is defined as "being feasible within an 8-h working shift". There are several commercial Food and Drug Administration-approved assays available which can detect AMR genes from a variety of specimen types. Progress has been slow due to a range of reasons including cost and regulation. Genotypic AMR characterisation methods are, however, being increasingly used in combination with machine learning algorithms in research to help better predict phenotypic AMR from organism genotype.
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Optical techniques such as phase contrast microscopy in combination with single-cell analysis are another powerful method to monitor bacterial growth. In 2017, scientists from Uppsala University in Sweden published a method that applies principles of microfluidics and cell tracking, to monitor bacterial response to antibiotics in less than 30 minutes overall manipulation time. This invention was awarded the 8M£ Longitude Prize on AMR in 2024. Recently, this platform has been advanced by coupling microfluidic chip with optical tweezing in order to isolate bacteria with altered phenotype directly from the analytical matrix.
Rapid diagnostic methods have also been trialled as antimicrobial stewardship interventions to influence the healthcare drivers of AMR. Serum procalcitonin measurement has been shown to reduce mortality rate, antimicrobial consumption and antimicrobial-related side-effects in patients with respiratory infections, but impact on AMR has not yet been demonstrated. Similarly, point of care serum testing of the inflammatory biomarker C-reactive protein has been shown to influence antimicrobial prescribing rates in this patient cohort, but further research is required to demonstrate an effect on rates of AMR. Clinical investigation to rule out bacterial infections are often done for patients with pediatric acute respiratory infections. Currently it is unclear if rapid viral testing affects antibiotic use in children.
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Vaccines.
Vaccines are an essential part of the response to reduce AMR as they prevent infections, reduce the use and overuse of antimicrobials, and slow the emergence and spread of drug-resistant pathogens.
Microorganisms usually do not develop resistance to vaccines because vaccines reduce the spread of the infection and target the pathogen in multiple ways in the same host and possibly in different ways between different hosts. Furthermore, if the use of vaccines increases, there is evidence that antibiotic resistant strains of pathogens will decrease; the need for antibiotics will naturally decrease as vaccines prevent infection before it occurs. A 2024 report by WHO finds that vaccines against 24 pathogens could reduce the number of antibiotics needed by 22% or 2.5 billion defined daily doses globally every year. If vaccines could be rolled out against all the evaluated pathogens, they could save a third of the hospital costs associated with AMR. Vaccinated people have fewer infections and are protected against potential complications from secondary infections that may need antimicrobial medicines or require admission to hospital. However, there are well documented cases of vaccine resistance, although these are usually much less of a problem than antimicrobial resistance.
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While theoretically promising, antistaphylococcal vaccines have shown limited efficacy, because of immunological variation between "Staphylococcus" species, and the limited duration of effectiveness of the antibodies produced. Development and testing of more effective vaccines is underway.
Two registrational trials have evaluated vaccine candidates in active immunization strategies against "S. aureus" infection. In a phase II trial, a bivalent vaccine of capsular proteins 5 & 8 was tested in 1804 hemodialysis patients with a primary fistula or synthetic graft vascular access. After 40 weeks following vaccination a protective effect was seen against "S. aureus" bacteremia, but not at 54 weeks following vaccination. Based on these results, a second trial was conducted which failed to show efficacy.
Merck tested V710, a vaccine targeting IsdB, in a blinded randomized trial in patients undergoing median sternotomy. The trial was terminated after a higher rate of multiorgan system failure–related deaths was found in the V710 recipients. Vaccine recipients who developed "S. aureus" infection were five times more likely to die than control recipients who developed "S. aureus" infection.
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Numerous investigators have suggested that a multiple-antigen vaccine would be more effective, but a lack of biomarkers defining human protective immunity keep these proposals in the logical, but strictly hypothetical arena.
Antibody therapy.
Antibodies are promising against antimicrobial resistance. Monoclonal antibodies (mAbs) target bacterial virulence factors, aiding in bacterial destruction through various mechanisms. Three FDA-approved antibodies target "B. anthracis" and "C. difficile" toxins. Innovative strategies include DSTA4637S, an antibody-antibiotic conjugate, and MEDI13902, a bispecific antibody targeting Pseudomonas aeruginosa components.
Alternating therapy.
Alternating therapy is a proposed method in which two or three antibiotics are taken in a rotation versus taking just one antibiotic such that bacteria resistant to one antibiotic are killed when the next antibiotic is taken. Studies have found that this method reduces the rate at which antibiotic resistant bacteria emerge in vitro relative to a single drug for the entire duration.
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Studies have found that bacteria that evolve antibiotic resistance towards one group of antibiotic may become more sensitive to others. This phenomenon can be used to select against resistant bacteria using an approach termed collateral sensitivity cycling, which has recently been found to be relevant in developing treatment strategies for chronic infections caused by "Pseudomonas aeruginosa". Despite its promise, large-scale clinical and experimental studies revealed limited evidence of susceptibility to antibiotic cycling across various pathogens.
Development of new drugs.
Since the discovery of antibiotics, research and development (R&D) efforts have provided new drugs in time to treat bacteria that became resistant to older antibiotics, but in the 2000s there has been concern that development has slowed enough that seriously ill people may run out of treatment options. Another concern is that practitioners may become reluctant to perform routine surgeries because of the increased risk of harmful infection. Backup treatments can have serious side-effects; for example, antibiotics like aminoglycosides (such as amikacin, gentamicin, kanamycin, streptomycin, etc.) used for the treatment of drug-resistant tuberculosis and cystic fibrosis can cause respiratory disorders, deafness and kidney failure.
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