Bench Press

Influenza viruses are negative-stranded, enveloped orthomyxoviruses that contain eight gene segments encoding various viral proteins. During the viral life cycle point mutations drive genetic drift responsible for seasonal influenza epidemics. Novel influenza viruses can also be produced through genetic reassortment between viruses. These viruses can result in devastating pandemics as they expose their host populations to novel antigenicity. These alterations in the influenza viruses’ genomes require that vaccine strains be updated annually to account for changes in virus populations.

Currently two types of vaccines are utilized to combat seasonal flu, each with their own limitations:

1. Chemically inactivated virus delivered via injection – mainly acts by inducing antibody response rather than cellular immunity and has suboptimal efficacy in elderly patients.
2. Live attenuated influenza virus vaccine of cold-adapted virus delivered via nasal spray – induces both humoral and cellular immunity, but is restricted to healthy children, adolescents, and adults, performing better in immunologically naive young children than adults.

So despite the relative effectiveness of the current offerings there is definitely room for improvement. That’s where research being done by Dr. Steffen Mueller and colleagues at Stony Brook University comes in. Building on prior research on producing synthetically attenuated polio viruses, Dr. Mueller’s group utilized a technique dubbed Synthetic Attenuated Virus Engineering (SAVE) to generate synthetic influenza virus vaccine candidates. They detail their work in a paper in Nature Biotechnology titled Live attenuated influenza virus vaccines by computer-aided rational design.

What’s truly unique about SAVE is that it only uses silent mutations within the viral genome to produce live attenuated virus. As described in the paper:

The central idea of SAVE is to recode and synthesize a viral genome in a way that perfectly preserves the WT amino acid sequence, while rearranging existing synonymous codons to create a suboptimal arrangement of pairs of codons. For reasons that are not understood, some pairs of codons occur more frequently, and others less frequently, than expected. … Although the mechanism of attenuation is unclear, preliminary evidence suggests that translation is affected. Attenuation can be ‘titrated’ by adjusting the extent of codon-pair deoptimization. Because codon-pair deoptimization results from miniscule effects at each of hundreds or thousands of nucleotide mutations (without changing amino acid sequences), reversion to virulence is extremely unlikely. Aided by computer algorithms, codon pair–deoptimized viral genomes can be rapidly designed and synthesized, and live virus can be generated by reverse genetics.

Figure 1

Utilizing SAVE Dr. Meuller’s group generated influenza strains with the following proteins synthetically deoptimized, polymerase subunit B1 (PB1), nucleoprotein (NP), and hemagglutinin (HA). They tested strains with a single deoptimized gene as well as one strain with all three altered genes. First Dr. Meuller’s group analyzed each of their strains in vitro to assess their growth characteristics. They found that all the mutant viruses produced plaques similar to wild type and produced reasonable titers although slightly lower (about tenfold lower). What was most interesting about their in vitro characterization however was their western blot analysis of the synthetically designed proteins. In each mutant it is clear that the gene that underwent SAVE produces significantly less protein in comparison to wild type (PR8) (Figure 1). The effect is specific and lends support to the idea that codon-pair deoptimization results in an effect on translation.

After checking their in vitro characteristics, Dr. Meuller’s group tested for attenuation by infected mice. They found that despite the reasonably robust viral growth each mutant strain demonstrated an attenuation effect. A strain combining all three modified genes (PR8^3F) resulted in an increase in median LD₅₀ of about 13,000 fold. Further tests on PR8^3F showed effective attenuation of symptoms and viral load in comparison to wild type (Figure 2).

Figure 2

In addition to assessing attenuation of the virus, Dr. Meuller’s group assessed the immune response and protective immunity in detail by immunizing mice and then challenging them 28 days after inoculation. As seen in Figure 3 below (a, b) strain PR8^3F had a much larger range of safe doses in comparison to wild type. Viral load is effectively limited (c) in PR8^3F inoculated animals and antibody response is robust (d) even in very low doses in comparison to LD₅₀ of PR8^3F.

Figure 3

Ultimately, this paper illustrates a really interesting new technology that seems capable of revolutionizing vaccine development. It provides a novel vaccine design strategy that appears to produce robust immune protection. It remains to be seen if SAVE vaccine candidates can address the limitations of current flu vaccines as well as other issues, but this paper is a strong first step for this technology.

(Source – Nature Biotechnology)

What would you do you if you were curious about the relative importance of selective pressures on a population of lizards on Caribbean islands? Since you’re reading Bench Press you might be inclined to turn to the power of computer modeling which can provide numerous advantages particularly when traditional experiments can’t be conducted. We’ve seen examples of computer models analyzing near earth asteroids, potential epidemics, and classic math and physics problems. However we’ve also seen that at times purely mathematical approaches can result in errors as well. Our inability to accurately describe problems with numbers 100% of the time makes it imperative that we continue to think creatively about ways to design experiments to test hypotheses.

That’s why I was particularly impressed by a paper in Nature by Ryan Calsbeek and Robert M. Cox, who wanted to explore the importance of selective pressures on anole lizards in the Caribbean. Field experiments to measure the effect of selective pressures are rare for a variety of reasons. A major one being the difficulty of finding animals and environments which can be manipulated in a controlled manner. Drs. Calsbeek and Cox didn’t let this stop them as they utilized a group of small islands in the Caribbean, each small enough to throw a ball end to end, as their test beds. There they removed the resident brown anole lizards and replaced them with experimental animals which had been carefully measured, tested for stamina, and tagged to identify at the end of the experiment.

Now that they had their experimental populations they needed to set up islands that tested the hypothesis that competition played a larger selective role than predation in island anole lizard populations. They established islands that had low and high density populations, and for each density type they setup islands inhabited by lizard-eating birds alone, lizard-eating birds and snakes, as well as islands free of predators (accomplished with a generous covering of netting as seen below). An unmodified control island was also monitored as a natural reference population.

They distributed the lizards in May and four months later at the end of the breeding season, September, they came back to capture the survivors and census the population. While this was difficult work they were able to collect a large amount of data which confirmed the hypothesis that competition is a more powerful selective force in these populations. They saw no real phenotypic differences in the lizards on islands experiencing differing predation, but saw that lizards surviving on crowded islands were significantly bigger and had greater stamina than those on less crowded islands (seen below). This indicated competition between lizards pushed the population while predation did not.  

While their clever experiment does a great job explaining the relative importance of selective pressures on this particular species of lizards on islands in the Caribbean it may not say anything about natural selection in other species. Despite that this paper remains awesome because as much as I like to see technology change the way we do science, I still appreciate a well constructed experiment to answer tough questions.

(Nature: Experimentally assessing the relative importance of predation and competition as agents of selection)

Having spent a few years working on cell based assays for screening small molecules I became aware of how limited traditional in vitro cell culture can be in modeling biological systems. Traditional tissue culture while fairly easy to do and manipulate for experiments, often produces two-dimensional growth with gene expression, signaling, and morphology that can be dramatically different from those found in vivo. This can make in vitro studies clinically irrelevant. In vivo work while a more accurate model has it’s own drawbacks such as cost and ease of manipulation. Therefore, it would be ideal to develop methods which can make in vitro tissue culture produce in vivo results.

That aim is what makes this paper by Souza et al. in Nature Nanotechnology so impressive to me. In this paper they describe a method for culturing cells three-dimensionally by magnetically levitating cells grown in the presence of a hydrogel consisting of gold, magnetic iron oxide nanoparticles, and filamentous bacteriophage. 

Dr. Souza’s group tested their hydrogel with glioblastoma cells as seen in the figure above. Application of a magnetic field allows the cells to counteract gravity floating in the media and allowing for three-dimensional growth. The field also concentrated cells resulting in cell to cell interactions consistent with previous work on tissue engineering scaffolds designed to provide a cell growth advantage. In addition, the shape of the magnetic field can also be used to shape cell growth.

While the ability to promote three-dimensional growth without biodegradable porous scaffolds or protein matrixes is remarkable, the truly impressive part of this technique is that the cells exhibit differential protein expression that more closely resembles that of in vivo tumor xenografts as seen in the figure below thanks to their new growth conditions.
The ability of these three-dimensional cultures to mimic in vivo samples effectively is remarkable and the simplicity of this technology could provide a less time intensive and cost effective solution to traditional experimental methods. It’d certainly be nice to someday be able to design in vitro assays which produce truly clinically relevant data. Maybe with techniques like this one we’ll be able to accomplish that soon.

(Source – Nature Nanotechnology : Three-dimensional tissue culture based on magnetic cell levitation)

Here at Bench Press we’re always interested in new initiatives that harness the advantages of the internet. We’ve covered various powerful distributive computing initiatives as well as breakthrough collaborative endeavors in scientific research. So I was intrigued when I saw buzz on Twitter about the Obama administration’s attempt to crowd source suggestions for scientific policy.

Through the American Association for the Advancement of Science (AAAS) and associated non-profit Expert Labs, the Obama administration wants to hear what grand challenges scientists envision taking on.

Expert Labs has a nice video explaining the reasoning behind this grand experiment in policy crowd sourcing.

After a quick search on Twitter I’m a bit curious as to how Expert Labs plans to parse all the data they’re going to get from this call to arms, but I’m optimistic that some interesting insights can be gleaned as to the opinions of Americans on the directions science should be headed in. More data never hurt right? If you’re interested in submitting an idea follow the directions here, you’ve got until April 15th!

Previously at Bench Press we’ve written about the power of distributive computing and it’s ability to pool resources from volunteers over the internet to tackle projects on protein folding and the search for extraterrestrial intelligence. As distributed computing approaches mainstream, numerous projects focusing on a variety of questions have emerged.

One project that caught my eye is the Quake-Catcher Network (QCN). The network is described as

a collaborative initiative for developing the world’s largest, low-cost strong-motion seismic network by utilizing sensors in and attached to internet-connected computers.

The QCN utilizes accelerometers attached to computers to monitor for vibrations. Vibrations detected by the accelerometer are then recorded and compared to readings from other computers in the network. Only when a sufficient number of computers report comparable readings at the same time will the data be reported as an earthquake. Most recently the QCN detected a magnitude 4.4 earthquake in the Los Angeles area yesterday morning. The data generated from QCN participants can be seen here.

The beauty of the QCN is the enormous cost savings their approach can provide in comparison to traditional seismic networks like those run by the USGS. New accelerometers are now much more affordable. Sensors that plug into a USB port can cost less than $50. In addition, an increasingly common feature for laptops is a built in accelerometer to detect sudden movements like drops in order to shut down components to protect them from damage. These accelerometers can be utilized and provide a fairly large potential participant base who merely need to install BOINC and join the QCN project to begin providing data to the network.

As the number of QCN participants grows the heads of the project, Drs. Elizabeth Cochran and Jesse Lawrence, hope the network will provide not only a wealth of data for geologists but potentially a small bit of warning in the event of a large earthquake for those miles away from the epicenter. Currently, Drs. Cochran and Lawrence are working hard to increase the number of participants while also providing educational tools for use in schools to teach about earthquakes and science behind them.

Living in San Diego I think I’m in a prime location to help out so I look forward to contributing some data to the QCN (magnitude 4.0 or less please!).

(Join the Quake-Catcher Network)(Source – LA Times)

We have always sought out new frontiers and this generation is no different. Today’s frontiers can’t be found on a map. They’re being explored in our classrooms and our laboratories, in our start-ups and our factories. And today’s pioneers are not traveling to some far flung place. These pioneers are all around us — the entrepreneurs and the inventors, the researchers, the engineers — helping to lead us into the future, just as they have in the past. This is the nation that has led the world for two centuries in the pursuit of discovery.

-President Barack Obama,  Energy Speech at MIT October 23, 2009

As a candidate for President and throughout President Obama’s first year he has established himself as a friend of the scientific community. He has made many statements like the one above poignantly establishing the importance of innovation and scientific discovery in our nation’s goals. However, supportive statements don’t pay the bills and as I’ve had reiterated to me several times during my conversations with various professors at graduate school interviews; funding is king. Ultimately, the best way for President Obama to demonstrate his support of the sciences is to show us the money. He started off on the right foot with the science funding handed out in the stimulus package last year. The chart below illustrates President Obama’s proposed budgetary changes.

Minus the small decrease in funding for the CDC, many scientific organizations could potentially see sizable increases in funding. As I’ll be starting my graduate work in biosciences next fall I’m extremely happy to see the proposed increases. I’m quite surprised to see a reduction in funds budgeted to the CDC after the recent swine flu outbreak. However, the overall increase in funding is a huge step forward compared to the prior administration. Hopefully Congress will pass a budget with increases similar to the President’s proposed budget. These funds could be used to improve the financial status of many labs across the country that were affected by declining funding availability. I’ve got my fingers crossed that President Obama will continue supporting the sciences in not just his speeches but in his actions as well.

(Chart – Wired.com)

We’ve written a couple of times about the asteroid Apophis which, while unlikely to hit Earth, will make a very near pass of 18,300 miles above the planet’s surface in 2029. NASA’s Jet Propulsion Laboratory just released an excellent animation of just how close Apophis will be when it passes by.

Wow for some reason 18,300 miles doesn’t seem quite as far anymore.
(Video Credit – Wired Science)

Here at Bench Press we’re fans of PLoS because they strive to expand access to the world’s scientific and medical literature with their open access stance as well as other experimental endeavors such as PLoS Currents: Influenza. That’s why when I checked in on PLoS Biology I was intrigued by a new collection titled Genomics of Emerging Infectious Diseases.

The collection is a series of essays, perspectives, and reviews discussing the potential genomics research holds in understanding emerging infectious diseases. While I haven’t had a chance to read through very much of the collection yet, one perspective written by Rajesh Gupta, Mark H. Michalski, and Frank R. Rijsberman suggests an interesting plan for infectious disease research. They suggest beginning an Infectious Disease Genomics Project (IDGP), much like the Human Genome Project.

The IDGP would be:

a coordinated, large-scale, international effort focused on the genomes of pathogens, vectors, hosts, and reservoirs and linked to end-point surveillance and response systems. Such a project could coordinate activities in four specific areas: generating data, linking data, analyzing data, and applying data.

The figure above illustrates some of the specific things the authors envision the IDGP being able to coordinate. Ideally the IDGP provides:

• A “roadmap” for researchers to follow in sequencing and monitoring emerging pathogens that allow researchers worldwide to aid in global efforts while continuing critical research on local diseases.
• Advanced data management in an easy to use, open-source, real-time interface. With an emphasis on linking as much data with relevant details (e.g. literature references).
• A centralized analytical toolbox with dynamic databases allowing for collaboration worldwide in addition to improved access for researchers in resource-limited settings.
• Ability to incorporate emerging technologies and provide access (e.g. new assay methods, next generation sequencers).

Personally I find the IDGP very intriguing simply from the standpoint of developing a framework for worldwide scientific collaboration. If this were successful it could provide a model for future projects. On a practical level, providing a network of this sort for scientists to rely on could at least increase the speed at which emerging diseases are detected. Increasing the speed of detection is always a good thing when dealing with novel pathogens with pandemic potential. It’ll be interesting to see what the scientific community thinks about beginning an IDGP.

Readers any thoughts?

(Source – PLoS Biology: Can an Infectious Disease Genomics Project Predict and Prevent the Next Pandemic?)

Previously we’ve covered the extensive tracking and modeling of near-earth objects NASA undertakes as well as efforts to pass along data to the public via the internet. In Ben’s post about modeling near-earth objects he wrote about a specific asteroid designated 99942 Apophis. Discovered in 2004, it has been closely scrutinized by astronomers worldwide as it’s size and potential for collision with Earth have sparked interest.

Meet Apophis. Discovered in 2004, it will likely set a record for harmless near earth pass in 2029.

Earlier data pointed to the asteroid potentially passing through a troublesome gravitational keyhole which increased the threat to Earth in 2036, however new data from previously unreleased images from a University of Hawaii telescope near the summit of Mauna Kea have allowed NASA scientists to improve their models. New models show a reduced risk of collision in 2036 from 1 in 45,000 to 1 in 250,000.

From the NASA press release:

“The refined orbital determination further reinforces that Apophis is an asteroid we can look to as an opportunity for exciting science and not something that should be feared,” said Don Yeomans, manager of the Near-Earth Object Program Office at JPL.

Modeling asteroids and allowing humanity a chance to risk assess is only one example of the power of computer modeling. However, this example also illustrates one important caveat about modeling. One’s model is only as good as the data utilized in generating and analyzing it. Let’s hope that future data on Apophis continues to produce good news.

(Image Credit)

What can you do with a 110 foot wide, nine story tall radio telescope that weighs almost a million pounds? Not a question the average person or even educational institution asks themselves. Yet, this was a question Lewis Center founder Rick Piercy contemplated when he convinced NASA to donate a radio telescope being decommissioned that had once been used to communicate with Apollo spacecraft.

Thanks to Rick Piercy’s efforts K-12 students around the world have access to the Goldstone Apple Valley Radio Telescope through the Lewis Center’s GAVRT program which is a partnership with NASA offering a variety of programs exposing students to radio astronomy and cutting edge scientific work. Teachers from all institutions are welcome to join the program and are given a training seminar and visit to the telescope in order to familiarize themselves with the curriculum currently designed around the GAVRT as well as learn how to control the telescope via the internet.

In one of the ongoing projects students have been giving NASA scientists a helping hand track the Lunar CRater Observation and Sensing Satellite (LCROSS) spacecraft with the GAVRT. The LCROSS mission is ongoing and the satellite is scheduled to make impact with the moon in order to look for water on October 9, 2009. One of the beauties of this program is that not only is this a unique learning opportunity for the students, but they also help provide additional data gathering time for NASA scientists as explained by Brian Day of the NASA Ames Research Center,

Because LCROSS has a very steeply inclined orbit, we have only a 2-hour window once every 3 days when we can check out the spacecraft using the Deep Space Network. So we decided to ask GAVRT for help. These kids help us get extra listening time for our spacecraft, and they get an incredible educational experience in return.

Thanks to the internet and some enterprising individuals some lucky students will have a front row seat to some exciting scientific exploration of our nearest celestial neighbor. I look forward to hearing about the results of the LCROSS mission. Congratulations guys!

Read more about GAVRT and the LCROSS mission.