Science Student

Would anyone like to contribute to Science Student?

Would anyone like to contribute to Science Student?

Nate

Pharmacofiles Has Moved

Pharmacofiles has moved to pharmacofiles.blogspot.com. My first post is a re-post of my introduction to the blog, so those of you who may have followed the blog from the beginning may want to skip the post. New posts will start appearing on Friday with the same triweekly schedule. So, coming up on Friday on Blogger is “Holy Fire.”

Thanks,

Neil  

Pharmacofiles is Moving

I hope everyone who enjoys this blog will join me at my new site. It will be hosted by google, but I do not have an address yet. Check back tomorrow for my new location.

Neil

Resistance Building

A battle is being waged between scientists and microbes. The scientists, with their laboratories, experiments, and intellect, are being matched blow-for-blow by microbes, single-celled organisms with a simple yet highly matable genome. Spontaneous DNA mutations have allowed microbes to keep one step ahead of medical science in the fight for survival. A recent reconnaissance of a microbial defense system published in the European Molecular Biology Organization Reports may help in the development of new weapons against the bacterial scourge.

The team of scientists crystallized two forms of the enzyme aminoglycoside 60-N-acetyltransferase. These enzymes inactivate antibiotics such as aminoglycosides and fluoroquinolones by transferring an acetyl chemical group to the antibiotic. The enzyme is responsible for broad-spectrum resistance that is it protects both gram-positive and gram-negative bacteria. Crystalography reveals the structure of proteins right down to the amino acid sequence. The active site of the enzyme, the site where the drug binds and a chemical reaction occurs is covered by a lid that forms a pocket. The pocket is formed so that bulky molecules such as aminoglycosides fit snugly inside. The amino acid sequence in the active site is variable, and this would explain how the enzyme can recognize many types of antibiotics, and how the resistance develops.

 Now that scientists know the physical structure and amino acid sequences of the enzyme, antibiotics can be synthesized that overcome resistance. One strategy would be to create antibiotics that are physically incapable of fitting into the pocket. Another approach would be to prevent acetylation after the antibiotic is bound. Any advance in our understanding of microbial defense systems puts us one step closer to overcoming resistance. 

Reference Maurice F, Broutin I, Podglajen I, et al. Plasticity and the emergence of broad-spectrum antibiotic resistance. EMBO Rep, 2008; doi:10.1038/embor.2008.9 

Coming Up On Monday…On the Move

Microbial Defenses

My post two days ago served as an introduction to antibiotics and the problem of microbial resistance to treatment with antibiotics. Today’s post takes the next logical step and describes the various mechanisms that bacteria use to resist our antibiotic attack.

Antibiotic resistance arises by mutation of bacterial DNA. The bacteria can transfer resistance to other bacteria either horizontally or vertically. Vertical transmission occurs when the bacterium splits and DNA is inherited by the daughter cell. Horizontal transmission occurs when the DNA is exchanged among bacteria by the processes of transformation, transduction, or conjugation. Transformation occurs when a bacterium takes up a naked piece of DNA from the environment. Transduction occurs when a virus carries the genetic material from one bacterium to another, and conjugation occurs when a bacterium forms a cytoplasmic bridge connecting to another bacterium through which the DNA passes.

 Mutated DNA encodes for proteins that differ from their native forms. Antibiotics that interacted with the native form of the protein may not be able to interact with the mutated form. These new proteins influence antibiotic action in several ways:

Some bacteria inactivate drugs, or do not activate them. The former method is used to inactivate aminoglycoside antibiotics and to inactivate penicillin. In the case of penicillin, some bacteria produce an enzyme called beta-lactamase, which destroys the lactam ring of the drug rendering it impotent. Some antibiotics require activation by bacteria before they have an effect. Tuberculosis treatment is complicated because of the failure to activate the antibiotic isoniazid.

Sometimes the antibiotic fails to reach its target. Pores and carrier proteins in the cell membranes of bacteria facilitate the entry of antibiotics. These entry proteins may be absent or mutated in resistant bacteria. Tetracycline and some other antibiotics are actually pumped out of the bacteria by efflux pump proteins. Resistance to methicillin occurs because some bacteria produce a protein that binds to the drug and prevents it from reaching its cellular target.

Finally, in some bacteria the protein that the antibiotic acts upon is mutated, as is the case for fluoroquinolone resistance.

 Recognizing these mechanisms is a vital step in overcoming resistance. For instance, a drug has been developed that inactivates beta-lactamase, the enzyme responsible for penicillin resistance. The next post describes a study designed to help this battle. 

Coming Up On Friday…A Reconnaisance Report on Bacterial Resistance

Microbial Strike and Counterstrike

In the late 1920s, a quiet and reserved scientist made one of the greatest breakthroughs in medicine. Alexander Fleming had been anxious to leave his laboratory behind and start his vacation. He was never very clean, and left behind some dirty Petri dishes on a desk next to an open window. When he returned, he found that the Staphylococcus bacteria left in the dishes had disappeared in areas where mold had grown. A laboratory below Fleming’s was experimenting with the Peniciiium notatum mold, which had become airborne and floated into the laboratory above. Eventually the antimicrobial secretion from the mold was identified and dubbed penicillin. The impact of penicillins on the treatment of infections was only realized many years later because Fleming understated his discovery.

The first antibiotic, penicillin and many related antibiotics such as the cephalosporins are not perfect. They are only effective against gram-positive bacteria such as Staphylococcus. Bacteria are classified as either gram-positive or gram-negative. Gram-positive means the bacteria will take up a dye and stain violet on a microscope slide. Gram-negative bacteria do not stain. Broad-spectrum antibiotics were identified that could kill both gram-positive and gram-negative bacteria. But a much larger problem has emerged that has stymied the battle against bacterial infections.

 The problem of antimicrobial resistance to antibiotics cannot be understated.  In the United States, more than 70% of infections acquired in the hospital are resistant to antibiotics that would have previously killed them. Many species of enterococci, Pseudomonas, and Enterobacter are resistant to all antibiotics identified to date.  The antibiotic vancomycin had been used sparingly because bacterial resistance to it was rare. Now vancomycin-resistant enterococci are common. Bacterial resistance occurs when the concentration of antibiotic needed to eradicate an infection exceeds the concentration that is not harmful to normal cells. The mechanisms of this resistance is the subject of my next entry. 

Coming Up On Wednesday…Weapons of Resistance

Tetrodotoxin: Of Zombies and Puffer Fish

Tetrodotoxin is the stuff of magic. It causes paralysis, anesthesia, loss of speech, but not loss of consciousness. It slows heart rate and respiration to levels that are barely detectable. Thus, poisoned victims are often reported to be in a zombie-like trance, or having risen from the dead once heartbeat and respiration return to normal. If enough poison is consumed, however, death usually results from paralysis of the respiratory muscles. Given these properties, tetrodotoxin is reportedly the major ingredient in the potions of witch doctors and used in voodoo ceremonies. Wes Craven’s film The Serpent and the Rainbow tells the story of a pharmaceutical representative who travels to
Haiti to investigate such a potion and determine if the ingredients could be used as an anesthetic. The film is based very loosely on the non-fiction book with the same name.

Tetrodotoxin is the stuff of pleasure. It is the poison in the puffer fish, also called balloon fish, blow fish, or fugu in
Japan. The puffer fish is so-named because it sucks in air or water to enlarge its throat when threatened. The toxin is secreted by bacteria, and is concentrated in the liver, gonads, intestines and skin. The term “tetrodotoxin” is derived from the Latin tetrodontis, or “four teeth.” In Japan and
Singapore, the fish is a delicacy. Specially-trained chefs prepare the fish. Properly-prepared fish should produce slight tingling of the tongue and mouth. Poorly-prepared fish cause death. About 150 accidental poisonings occur in
Japan each year, and about half of the victims die. Some newts, frogs and octopi also produce the toxin.

Tetrodotoxin is a channel blocker. It blocks voltage-sensitive sodium channels on cell membranes, which prevents sodium entry. In this way the toxin prevents conduction of nerve impulses. The toxin is too dangerous to use therapeutically, but it is a useful experimental tool. Tetrodotoxin, for example, can be used to determine how drugs influence the nervous system.

 Tetrodotoxin poisoning is treatable. The drug neostigmine has been reported to help prevent death from tetrodotoxin poisoning. The drug inhibits the enzyme acetylcholinesterase, an enzyme that would normally break down the neurotransmitter acetylcholine. The accumulation of acetylcholine can help stimulate respiratory muscles and prevent respiratory failure. 

Coming Up On Monday…An Unwinnable Fight?

Not Quite Placebos

Orson Welles played one of the most despicable villains in the history of films. In The Third Man, Welles portrayed a racketeer named Harry Lime. The plot detailed the efforts of a novelist, played by Joseph Cotton, to clear the “good” name of his friend Lime. In the process, however, Cotton discovered his friend was not as he appeared to be. In one of many memorable scenes, Cotton is shown children who were treated with penicillin that Welles supplied. The children were suffering horribly because Welles was peddling poisonous penicillin. We never actually see the children. Rather, our imaginations take over as we listen to the physician’s descriptions of the children and watch Cotton’s reactions. The film is a classic of cinematography, writing, directing, and acting, and must be seen. Even and episode of the television show Law and Order featured a courtroom showdown in which the DA described the famous Ferris wheel sequence in the movie. The episode, like the movie, described the selling of drugs that contained little to no medicinal ingredients. No, they were not peddling homeopathic “cures.” These “drugs” were sold with the names of legitimate medicines. A recent paper in the Public Library of Science: Medicine journal investigated these fake drugs in an effort to track down the counterfeiters.

Since 1998, southeast Asia has seen an explosion in the amount of counterfeit artesunate. Artesunate is a drug used to treat malaria in
Africa and southeast Asia. Health officials, government agencies, and Interpol collaborated on the collection of fake and genuine artesunate samples from several southeast countries. The chemical composition of the fake artesunate was determined, which would aid in finding the source of the fakes. The packaging of some fake drugs was very similar to the packaging of authentic medicine, whereas some packaging of fakes was fairly crude. Close scrutiny would be needed to identify packaging of some of the fraudulent medicine. The cruder packaging contained spelling errors, font inconsistencies, and other obvious markers of fraud. Half the confiscated drugs were suspected of being fakes based on packaging, and chemical analysis confirmed that these drugs were indeed fake. The fake drugs contained little to no artesunate, but rather low amounts of acetaminophen (Tylenol), some antibiotics (erythromycin, metronidazole, chloramphenicol), the antimalarial chloroquine, and other chemicals. Some of the drugs contained the carcinogens benzene and safrole. Some samples even contained high amounts of charcoal. This is probably because some of the drugs are obtained from plants, and these plants were grown in highly-polluted areas.

 The investigators used the chemical composition of the fakes to identify where the drugs were being made, and several arrests were made. The authors point out that the low amount of antibiotics and genuine artesunate found in some of the fakes could contribute to the growing problem of antibiotic resistance. And I would add, just like in The Third Man, these fake drugs are causing suffering and death.  

ReferenceNewton PN, Ferna´ndez FM, Planc¸on A, et al. (2008) A collaborative epidemiological investigation into the criminal fake artesunate trade in South East Asia. PLoS Med 5

Coming Up On Friday…The “Four teeth” toxin