February 25, 2010 at 4:05 am #2805
The video interview is complete, see the link within the next post. Also, sink your teeth and neurons into the attached article and transcript excerpt!
Engineered bacteriophage targeting gene networks
as adjuvants for antibiotic therapy
Timothy K. Lu and James J. Collins
Harvard-Massachusetts Institute of Technology Division of Health Sciences and Technology, Cambridge, MA 02139; and Howard Hughes Medical Institute, Center for BioDynamics and Department of Biomedical Engineering, Boston University, Boston, MA 02215
Bacteriophage Platforms to Overcome Antibiotic Resistance
Bacterial infections affect hundreds of thousands of people in the United States every year, and in some cases, they are serious, even fatal. Though scientists have developed a number of effective antibiotics to fight off harmful bacteria, many of these microscopic organisms have developed resistance to such medications. This, combined with the fact that developing antibiotic drugs is extremely expensive and resource-intensive for drug companies, means that the overall effectiveness of this class of medications could be poised to decline.
Synthetic biologist Timothy Lu aims to turn this issue around with his work on bacteriophage, viruses that attack only bacteria, not human cells. He hopes the results of his work will be effective at destroying some types of antibiotic-resistant bacteria. His inventions are aimed at helping to enable the rapid design and production of inexpensive antibacterial agents that have the power to boost antibiotic effectiveness.
Lu was born in 1980 in Stanford, California, and raised in Yorktown Heights, New York, and Taiwan. His interest in scientific endeavors was inspired in part by his father, an engineer-entrepreneur who has worked in the semiconductor and integrated circuits industries.
Lu pursued M.D. and Ph.D. degrees via the Harvard-MIT Division of Health Sciences and Technology program, a partnership among the Massachusetts Institute of Technology, Harvard University, Harvard Medical School and several Boston area teaching hospitals and research centers. While working at a hospital as part of a graduate course, he saw many patients who contracted new infections due to already-compromised immune systems or equipment that is extremely difficult to keep sterile. He identified this as a very serious problem and thought there must be something he could do to help.
With his advisor, J.J. Collins, a professor at Boston University, Lu began looking at bacteriophage as a possible platform for overcoming antibiotic resistance. While these viruses have been used for nearly a century to treat bacterial infections, their popularity declined among the medical community when antibiotics became readily available. Lu decided to apply synthetic engineering to develop bacteriophage specifically aimed at boosting antibiotic effectiveness. Simply explained, the bacteriophage carries DNA that codes for factors that target bacterial gene networks, which former treatments failed to reach, and destroys bacterial antibiotic resistance mechanisms. The weakened bacterial defenses enable antibiotics to perform better.
Administered together, Lus bacteriophage and antibiotics have the potential to eliminate nearly 30,000 times more bacteria than antibiotics alone, including cells that survive antibiotic-only treatment. This combination treatment also thwarts development of stronger antibiotic resistance, which can extend the lifetime of existing and future antibiotic drugs.
Lu has also applied his work with bacteriophage to create a new technique for reducing harmful biofilms, slimy layers of bacteria that develop on the surfaces of medical, industrial and food processing equipment and are difficult to penetrate and remove. Current treatment methods to penetrate biofilms can involve peptides or enzymes, which must be administered systemically and are costly. Medical devices infected by biofilms, such as replacement hip joints or pacemakers, often have to be removed surgically.
Lu invented enzymatically active bacteriophage that directly target the infection site, where they can simultaneously penetrate the biofilms protective slime layer and kill the bacteria underneath. In tests, his platform proved greater than 99.997 percent effective at destroying biofilms, a significant improvement over current treatment options.
In recognition of his efforts, Lu was awarded the 2008 Lemelson-MIT Student Prize.
PNAS Early Edition.pdf
March 24, 2010 at 9:32 pm #3362
Question: The studies from the CDC state that 60-80% of chronic infections involve biofilms. Whats the connection between bacterial biofilms and chronic health conditions?
Answer: Theres an increasing recognition that biofilms play a role in chronic diseases and the reason for that is that biofilms are very difficult to eradicate. They live on surfaces, they produce extracellular material that can defend themselves against human immune cells or antibiotics, and therefore theyre very difficult to remove. And once they form on a surface, theyre very difficult for your body or for drugs to kill them. Examples of this can include infections of bone, infections of your heart valves, infection of the urinary tract, in which bacteria can colonize these surfaces of the human body and live there for long periods of time.
Question: Tell us a bit more about biofilms.
Answer: So biofilms are a complex type of organizational structure that bacteria and these yeasts like to live in. I like to generally think of bacterial biofilms kind of like fruit Jell-O. Thats kind of the analogy I like to talk about, and the reason for that is, if you think about a fruit Jell-O, where you have the individual pieces of fruit, embedded in this gelatinous material, the biofilm cells, the bacterial cells, are like the fruit in the, inside the Jell-O and they produce materials, for example, polysaccharides, DNA, all those types of different materials that form this gelatinous layer that surrounds them and provides structure for them to live in. Depending on the species of the bacteria, if youre talking about E. coli or staphylococcus species or pseudomonas species. The type of extracellular material they produce is different, but they all have one thing in common: they provide some kind of protection and provide some stabilization structure for the rest of the community.
Question: How does a colony form, when does it form…what are the conditions necessary for a biofilm to form?
Answer: The life cycle of a bacterial biofilm is pretty interesting. So it would usually involve some type of bacteria that can swim around we call planktonic culture, which is in liquid, and theyre not stuck to any surfaces. They swim around until they find a surface that they like, and then they stick to that surface, and they start growing on that surface and growing more and more complex structures as they grow. At a certain point, when the biofilm is grown to a certain size, it reaches its mature state. At that point, it can shed bacteria that kind of can swim to other locations. This happens in human cells, the human body, happens out in the environment, and the conditions that are necessary for biofilm formation are essentially like a surface that the bacteria would like to live on and the bacteria needs to be able to produce the right types of material so he can stick to those types of surfaces.
Question: How do you think the biofilm colonies are hiding from the immune system and/or what makes a biofilm more resistant to interventional therapies?
Answer: Biofilms are difficult to eradicate types of infections for a variety of different reasons. is just by the purely that they live on surfaces. So, if you think about it, if you have a bacteria floating around in a liquid and your immune cell comes by and tries to eat it, thats very easy for the immune cell to reach around the bacteria and eat it, as opposed to if youre trying to clear something off of a surface, theres no easy way for your immune cells to kind of get around the infection. It cant, it cant really flank the type of infections. So thats one kind of big problem in terms of trying to eradicate biofilms from any surface.
The second reason is that when a biofilm forms, it tends to produce these extracellular materials that can either prevent the penetration of things like bacteriophages or immune cells deep into the biofilm.
The third thing is that when biofilms form, a lot of these cells adopt what is called a persistence state, in which they kind of shut down, theyre not very metabolically active, and, as a result, a lot of the drugs that we use, like antibiotics, cant really kill those cells. A lot of the antibiotics we use actually target actively dividing cells. So, if a cell is kind of just sitting around, not dividing, not very active, its very hard for antibiotics to kill these types of cells.
I think thats definitely a problem, since more research has shown that especially some of the antibiotics that are very active against killing bacteria can never sterilize an entire population. So lets say you start off with a culture of a million bacteria, and you apply this very strong antibiotic. You may kill off the majority of that bacteria, but, for example, some of the studies weve done show that, lets say you start off with a million bacteria, you may be able to kill 999,000 of them, but you still have about a thousand bacteria left, which are kind of immune to the antibiotics at this point. They can adopt a persistence state, and once you remove the antibiotics, they can start forming again and growing into a biofilm. So the presence of persister cells and the inability for us to really completely eradicate a biofilm leads to things like antibiotic resistance and future biofilm formation.
Question: How many different microbes work together to create biofilm communities? E.g., gram negative, gram positive, yeasts ?
Answer: I think your question about how different microbes work together to create biofilm communities is a very interesting one, and its something that the communities only now are really starting to get a grasp on. There are many theories, where how different microbes can work with each other and the limitation is that we dont necessarily have great technologies by which we can study these. There are several mechanisms by which this might work. For example, bacteria can produce these molecules called quorum sensing molecules, and they produce these when the bacteria grow to a significant concentration, and it kind of signals to their friends, like a bacterial cell will tell its friends, like, Oh, now that weve reached a certain concentration, maybe we should switch into a different state, like a biofilm state or a virulent state. And its been found that, actually, different types of bacteria can communicate with each other by producing these quorum sensing molecules, and so quorum sensing is a big component of what people are trying to study now in terms of polymicrobial biofilms.
In other situations, different microbes may have kind of more of a symbiotic relationship, where one microbe might produce a nutrient that the other one likes and vice versa, and therefore, they would like to form a, a biofilm where theyre in close proximity to each other, and they can both have increased survival because they live in a biofilm together.
The third thing maybe that different microbes produce is different factors that can help protect each other from extracellular insults. For example, bacteria may produce a certain type of polysaccharide that another one doesnt, and in these polymicrobial biofilms, they may produce different types of polysaccharides that add increased protection for the entire community as a whole. And the last thing that people like to talk about in terms of polymicrobial biofilms is that, because these different cells are in close proximity with each other. Theyre in an ideal situation where one bacterial species can transfer genes to another one, and they can share genetic elements that can promote survival. One of these survival elements are antibiotic resistance genes, for example, and that can be traded between bacteria, and cause antibiotic resistance to spread through a community of different types of species.
Gene transfer can occur between bacteria or between bacteria and non-bacterial species — studies have been done between bacteria — theres a huge diversity of bacteria that have antibiotic resistance and even just studying that reservoir of antibiotic resistance genes is quite interesting already.
Question: How are your T3 and T7 phages able to enter bacterial biofilms and is the hydrolysis of the biofilm adhesives the key part of that strategy? Is it akin to like chipping away the cement between bricks?
Answer: Thats a great analogy. So the way our engineered bacteriophage technology works against biofilms is that they produce components, or these enzymes that can chew up the biofilm, extracellular matrix. So, as I mentioned before we like to think about these things as fruit Jell-O, where the, the fruit inside of the biofilms are the cells and the Jell-O is the extracellular components. And, if you think about it, if youre trying to deliver an antibiotic or virus deep into the fruit Jell-O, its going to be very hard to penetrate through all the Jell-O to the bottom of the bowl. So, by producing enzymes that can chew up the gelatinous material you can allow these phages and antibiotics to penetrate deeper and deeper into the biofilms and eradicate much more of the biofilms than youre able to do otherwise.
Question: Regarding your future solutions: are you focusing on monomicrobial, or could it also be true for the polymicrobial films?
Answer: So our technology, as currently tested, is focused on the monomicrobial biofilms, and this is for a variety of reasons. For regulatory reasons, its advantageous to go after monomicrobial biofilms when youre trying to put a, push a new technology through with the FDA. Its also easier to work with in the laboratory. However, we do believe that a lot of these technologies are applicable to polymicrobial biofilms, and the reason is as follows: all the enzymes that were using actually target a variety of different types of bacterial biofilms. So, not only do they go after E. coli, but they go after other different types of bacteria as well, the same enzyme, and therefore we think that it should be able to go after polymicrobial biofilms as well, although its something that is a little bit further along in our research plan.
Question: Can you differentiate between acute conditions, chronic conditions, systemic bacterial infections and where your future solutions may apply?
Answer: So the technology that were developing can probably be used for acute conditions, chronic conditions, and in certain cases, systemic infections as well. Acute bacterial infections, such as the ones that cause sepsis, in which you have bacteria flowing through your bloodstream can be treated with bacteriophage, although it may not be the ideal technology to use at that point. In those acute infectious conditions, the bacteria usually has not had enough time to form a very thick biofilm, and therefore those bacteria generally can be killed by antibiotics, and these antibiotics can spread throughout your bloodstream very quickly, and can treat these types of infections.
Question: Are there any things, any other things you wanted to mention today that we didnt cover that are important for the audience to hear?
Answer: Biofilms are actually intricately related to the problem of antibiotic-resistant bacteria, which is a topic people have heard about a lot more in the popular press. So antibiotic-resistant bacteria are essentially infections that are difficult to treat because these bacteria have evolved resistance to antibiotics. But awareness of the biofilm issue has really spread significantly throughout the medical community.
Examples include MRSA infections, but there are a lot of other examples, e.g., gram-negative and gram-positive bacteria that have acquired genes that make them immune to a lot of the antibiotics weve developed. Now the issue is that, in the last 30 years, investment in developing new antibiotics has really decreased significantly, and, therefore clinicians are very restrained at this point in terms of what antibiotics are effective in terms of treating these types of infections.
The link between biofilms and antibiotic-resistant bacteria is quite close; in that biofilms, because of the presence of persister cells and being able to trade genes between each other, can often have an increased level of antibiotic resistance. And so moving forward, the type of technologies we need to develop and the type of research that we need to do should really focus on both of these problems together: biofilms as one distinct entity, but also on antibiotic resistance and how they interplay with each other to cause human disease.
April 19, 2010 at 4:43 pm #2806
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