The virus-antibody arms race

One of the new concepts I learned when I started working on HIV was the most recent common ancestor, or MRCA. When you look at the genetic make-up of a population, you will find a certain amount of variety but also a much greater amount of overlap, i.e. stretches of DNA that are identical throughout the population. Using phylogenetics, one can look at these patterns of shared vs. mutated stretches, and reconstruct the genetic ancestor of the population. For example, you've probably heard of Mitochondrial Eve: since we all inherit our mitochndrial DNA from our mothers, scientists have been able to look at the mitochondrial DNA across all populations and determine the one ancestor (our common mother, so to speak) from which they all originated. Pretty cool, right?

My line of work, for the past 6-7 years has been estimating most common recent ancestors, or MRCAs, of HIV-1 populations. A few years ago we found that in sexually transmitted infections only a handful of viruses are able to come across the genital mucosa and start the infection. Therefore, if you draw a blood sample early enough (a few weeks) after the start of the infection, from that sample we can infer the MRCA of the viral population in the patient. This is particularly relevant because in the case of a viral infection, the MRCA is likely to be the virus that initiated the infection. As the infection progresses, the viral population changes, but it is the ones that are able to break the mucosal barrier (i.e. the MRCAs) that a vaccine needs to target.

Once inside the host, viral evolution is (for the most part) driven by the host's immune system as it tries to counter-attack the infection. At the same time, as the virus changes its genetic make-up to escape the immune pressure, the immune system itself changes and tries to come up with new ways to neutralize the enemy. It's an arms race that in HIV infections typically sees the immune system always one step behind: the first antibodies found in an HIV-1 infected person react with the first, unmutated virus that initiated the infection (the MRCA). As the infection progresses and the virus evolves, new antibodies are made that are able to react to the following viral generations, but typically there's always a subpolulation of viruses that's one step ahead of the antibodies and can still escape. (I hope this part is clear, I've been struggling quite a bit to find the right wording for this paragraph, so if it's not clear feel free to ask questions in the comments.)

In order to design an efficient vaccine, we need to find a way to elicit broad neutralizing antibodies, where by "broad" we mean antibodies that react not only to the present or past viral generations in one host, but to a wide variety of viruses across different hosts and populations. Such antibodies are found in a minority of HIV-infected patients and, typically, by the time they arise, the infection is so spread that they cannot clear the virus.

Ideally, a vaccine should boost a "short-cut" in the evolutionary path that leads to the production of broadly neutralizing antibodies much faster than our bodies are currently capable of. Unfortunately, all vaccine trials attempted so far have not been able to elicit broad neutralizing antibodies. Why?

Antibodies are made by B-cells, white blood cells produced in the bone marrow. In order to produce antibodies, B cells need to be activated, which happens when they find an antigen specific to their receptor. Once activated, B cells not only start producing antibodies, but they also either become memory cells (so that if the antigen is encountered again, the immune system will know which antibodies to produce in order to clear it) or they undergo further differentiation. This process of undergoing more differentiations ensures that the "match" between receptor and antigen becomes tighter and tighter. It takes many cycles of differentiations to produce HIV-1 broadly neutralizing antibodies, and, currently, the process takes so long that most patients don't produce them ever, and the ones that do, don't get them in time to clear the infection.

One reason why we believe it takes many differentiations to make HIV broadly neutralizing antibodies is that they share many similarities to self-reacting antibodies, antibodies that are normally destroyed by the body because they carry a high risk to originate auto-immune disorders (when the immune system attacks its own self instead of antigens). So, instead of eliciting the actual antibodies, could a vaccine elicit its ancestor? Remember how I said that the viral population constantly evolves and, hand in hand, so do the antibodies? Since we can estimate the viral ancestors, can we do the same for the antibodies? Can we reconstruct the differentiation pathway that leads to broadly neutralizing antibodies?

In [1], Liao and colleagues have reconstructed the lineage of the infecting virus in one African HIV-infected patient (CH505), as well as the lineage of an antibody, found in the same patient, able to neutralize 55% of ~200 HIV-1 isolates. the researchers effectively reconstructed the coevolution of virus and antibody within the patient. The patient was followed from week 6 after the infection up until 236 weeks after the infection, and during this period no antiretroviral therapy was administered. This is important because it means that the viral evolution was driven solely by the immune pressure.

Liao et al. found that the first unmutated ancestor in the B-cell lineage appears at week 14 after the infection, and it keeps mutating in ways that are reflected in the evolution of the virus. Once they retraced all the intermediate steps that led to the production of the broadly neutralizing antibody, the researchers tested all of the intermediate antibodies for reactivity against the virus, from the infecting strain to its later generations. They found that breadth and strength of reactivity increased as the antibody lineage evolved. In light of what I tried to explain above, this is a fantastic step forward in understanding how the virus evolves under the immune pressure, as it can help design a vaccine that elicits antibodies that are one step ahead (instead of behind) in the virus-host arms race.

"Thus, a candidate vaccine concept could be to use the CH505 transmitted/founder Env or Env subunits (to avoid dominant Env non-neutralizing epitopes) to initially activate an appropriate naive B-cell response, followed by boosting with subsequently evolved CH505 Env variants either given in combination, to mimic the high diversity observed in vivo during affinity maturation, or in series, using vaccine immunogens specifically selected to trigger the appropriate maturation pathway by high-affinity binding to the unmutated common ancestor and antibody intermediates. [. . .] The finding that the transmitted/founder Env can be the stimulator of a potent BnAb and bind optimally to that broadly neutralizing antibody unmutated common ancestor is a crucial insight for vaccine design, and could allow the induction of broadly neutralizing antibodies by targeting unmutated common ancestors and intermediate ancestors of broadly neutralizing antibody clonal lineage trees."

Of course, there's the usual caveats: will this kind of pathway be reproducible in other patients? How much of it is randomness and how much is it not only retraceable but reproducible is something we will only understand by getting more data from more patients. But it's a start, and a very promising one.

[1] Liao, H., Lynch, R., Zhou, T., Gao, F., Alam, S., Boyd, S., Fire, A., Roskin, K., Schramm, C., Zhang, Z., Zhu, J., Shapiro, L., Becker, J., Benjamin, B., Blakesley, R., Bouffard, G., Brooks, S., Coleman, H., Dekhtyar, M., Gregory, M., Guan, X., Gupta, J., Han, J., Hargrove, A., Ho, S., Johnson, T., Legaspi, R., Lovett, S., Maduro, Q., Masiello, C., Maskeri, B., McDowell, J., Montemayor, C., Mullikin, J., Park, M., Riebow, N., Schandler, K., Schmidt, B., Sison, C., Stantripop, M., Thomas, J., Thomas, P., Vemulapalli, M., Young, A., Mullikin, J., Gnanakaran, S., Hraber, P., Wiehe, K., Kelsoe, G., Yang, G., Xia, S., Montefiori, D., Parks, R., Lloyd, K., Scearce, R., Soderberg, K., Cohen, M., Kamanga, G., Louder, M., Tran, L., Chen, Y., Cai, F., Chen, S., Moquin, S., Du, X., Joyce, M., Srivatsan, S., Zhang, B., Zheng, A., Shaw, G., Hahn, B., Kepler, T., Korber, B., Kwong, P., Mascola, J., & Haynes, B. (2013). Co-evolution of a broadly neutralizing HIV-1 antibody and founder virus Nature, 496 (7446), 469-476 DOI: 10.1038/nature12053

Wallflowers!

I'm thrilled to announce that four of my flowers will be displayed at the Fuller Lodge Art Gallery in Los Alamos, NM, from June 14 until July 27th in a collective titled "Wallflowers." Lots of other artists from NM will be there, so check-out the link in a few days and you'll be able to see a preview of their work. These are mine:

Can't make it to NM this summer? No worries, you get a second chance to see my work in the fall: yes, I will have my very first gallery show in Santa Fe, NM, this coming fall: from September 27 until October 8th my flowers and sunsets will be at the Silver Sun Gallery on Canyon Road, one of the most characteristic roads in Santa Fe. Hope you can make it then and stay a few more days for Balloon Fiesta in Albuquerque! :-)

Bug hunting and other NM encounters

Whenever I put the macro lens on my camera and set out for a field trip I learn something new. Last year I found a metallic green sweat bee and a yellow bee with blue eyes. This year I found a black bee with green eyes and ...

A yellow heart bug (yes, I know, none of these are scientific names, in fact, if somebody can provide a scientific name, please do so in the comments, I'll edit them in):

A golden bee:

And (drum roll, please, because I think this one's the star of the show) a metallic blue bee:

EDIT: thanks to Steven Halter's attentive eye, this beautiful bee has been identified as Osmia ribifloris. Thanks, Steve! Here she is, again, poking her head out of the thistle thorn and realizing she's no longer alone. "Ladies! I was here first!"

Bugs weren't the only encounters. This fellow gave us a lesson on where to properly cross the road. A huge SUV politely stopped and waited.

Finally, I did swap lenses at some point and took this picture for my friend Hollis, who's the expert on rock formations and their awesome colors. These are red rocks in Jemez Springs, NM.

Happy Memorial Day every one! :-)

Should you worry about vitamin D deficiency? Maybe. Or maybe not.

Since my last blog post, where I shared my thoughts on BRCA1, BRCA2, and preventive mastectomies, I've been asked what else can a woman do to reduce her risk of breast cancer. I've heard a big deal about vitamin D, so I did a bit of research on the matter.

As a disclaimer, I should tell you up front that, though many correlations between vitamin D deficiency and cancer risk have been found, just as many have been refuted or found inconclusive. You can read more about it on the wikipedia page.

What is vitamin D?

The name "vitamin D" includes a group of steroid-like molecules (they are similar to steroids, but not quite steroids) that help our intestine absorb calcium and phosphates. Since calcium is essential in bone development, vitamin D deficiency has been most commonly associated to osteoporosis and other bone-related diseases. There aren't many foods rich in vitamin D, however, vitamin D can be endogenously synthesized when the skin is exposed to sunlight. Unfortunately, modern lifestyle keeps us cooped up many hours in office cubicles, or in the house during chores, or in malls. When we're out enjoying the sunshine we cover up with hats and super-protective sunscreens because we've been told that the sun is bad for the skin and can cause malignancies. As a consequence, vitamin D deficiency is increasing world-wide.

There is a foundation for all the studies that have analyzed correlations between several diseases, including cancers, and vitamin D: (i) several ecological studies have found a trend for an increase in incidence of certain cancers at higher latitudes, suggesting that longer exposures to the sun may have a protective effect. (ii) The vitamin D receptor (VDR) is expressed in many cells of the immune system, and mouse models have shown that vitamin D deficiency can promote certain auto-immune diseases. In a recent review, Sundaram and Coleman examine the link between vitamin D and influenza [Adv. Nutr. 2012 3: 517-525]. (iii) "VDR regulates a wide range of cellular mechanisms central to cancer development, such as apoptosis (cell death), cell proliferation (uncontrolled cell growth), differentiation, angiogenesis, and metastasis [1]". In line with this observation, Pereira, Larriba, and Munoz published a review on the evidence that vitamin D plays a protective role in colon cancer [Endocr. Relat. Cancer 2012 19: R51-R71].

In [1], Crew discusses the use of vitamin D supplementation as part of breast cancer prevention. She presents many interesting findings, for example:

"Colon, breast, and lung cancer have all demonstrated downregulation of expression of VDR when compared to normal cells and well-differentiated tumors have shown comparably more VDR expression as measured by immunohistochemistry when compared to their poorly differentiated counterparts. Higher tumor VDR expression has also been correlated with better prognosis in cancer patients [1]."

Crew looks at different types of studies: some suggest beneficial effects from using vitamin D (calcitriol) in combination with other anti-cancer treatments; some found an inverse association with mammography density, a biomarker for breast cancer (supposedly high density increases the risk of cancer); some found an inverse association between better breast cancer prognosis and vitamin D deficiency. However, many of these studies have limitations. For example, some only assess the levels of vitamin D through dietary intake, which is not a good measure of the circulating levels because it doesn't account for vitamin D synthesized through sun exposure. Some were confounded by obesity since fat is known to sequestrate vitamin D and also raise breast cancer risk. In light of all these considerations, Crew concludes:

"Even with substantial literature on vitamin D and breast cancer, future studies need to focus on gaining a better understanding of the biologic effects of vitamin D in breast tissue. Despite compelling data from experimental and observational studies, there is still insufficient data from clinical trials to make recommendations for vitamin D supplementation for breast cancer prevention or treatment [1]."

As I often do in my posts, rather than giving you answers, I make an effort to provide you with pointers and food for thought: in the end you have to make your own decisions about your health and the wellbeing of your family. As a personal note, I'll add that on my last blood report my vitamin D circulating levels were undetectable. I had no symptoms whatsoever, but I am now taking a vitamin D supplement. I'm also much less paranoid about smothering my kiddos with sunscreen when they play outside (which has made them much happier, two birds with one stone).

[1] Crew, K. (2013). Vitamin D: Are We Ready to Supplement for Breast Cancer Prevention and Treatment? ISRN Oncology, 2013, 1-22 DOI: 10.1155/2013/483687

Angelina no longer has them. Does that mean I should get rid of them too?

We love them and yet we hate them. They get censored, augmented, reduced, replaced, covered, exposed. They get grilled, occasionally, but those are not the ones I'm talking about. We want to see them and yet we pretend we don't. We criticize them and yet we forget what they are made for, the most beautiful thing of all: nourish a new life.

Yes, I'm talking about breasts.

Angelina Jolie's breasts have been extensively discussed this week, more now that they are reportedly gone than when they were around. Sort of ironic, if you thin about it. Angelina did the unthinkable: she had both her healthy breasts removed to prevent cancer. In a second phase of her preventive plan, she will have her ovaries removed, too. The tabloids will no longer be able to speculate on her possible new pregnancies, but they will have plenty to discuss on and around her missing body parts.

Somehow the news left me a little puzzled, unable to share the views of those who praised Angelina for her bravery. Yes, it takes guts to do what she did. At the same time, the huge resonance she's been given seems blown out of proportion. Just another Hollywood thing. It reminds me of back when our mothers were told that formula was way better than breast milk. Are we facing a new era where silicon is better than milk ducts? Are they trying to convince us that fake is healthier than real? Well, of course it is. It's fake!

So, before we go around demonizing breasts and invoking chopping off body parts in the name of longevity, I wanted to get some facts straight.

First of all, I read over and over again, "Angelina Jolie carries the gene BRCA1 ..." Turns out, we all carry the gene. What makes us different is that there are distinct copies of this gene across individuals, and some copies (but not all) do raise the risk of breast and ovarian cancer.

BRCA1 and BRCA2 are part of the so called tumor suppressor genes, genes that code for proteins that are in charge of repairing damaged DNA. Our cells undergo numerous cellular divisions during our lifespan, and every cell division carries a certain chance of damaging the DNA. Though rare, mutations can be introduced, which can either be lethal or create a cancerous cell. Tumor suppressor proteins make a first attempt to repair the damaged DNA. If the DNA cannot be repaired, they promote apoptosis, or cell death. Another example of tumor suppressor gene is TP53, which encodes the protein p53.

The first link between BRCA1 and breast cancer was discovered in 1990 by Hall et al. [1]. BRCA1 and BRCA2 are expressed mostly in breast tissue. Some mutations in these genes cause them to code proteins that are not fully functional. When this happens, a cell with damaged DNA has a higher chance to escape the "screening" and start dividing instead of undergoing apoptosis. Because BRCA1 and BRCA2 are expressed mostly in the breast tissue, by removing the breast tissue one gets rid of the majority of cells expressing the defective genes, which in turns significantly lowers the chance of developing breast cancer.

While hundreds of mutations/variations in the BRCA1 and BRCA2 genes have been found, not all are linked to breast cancer, and the ones that are don't increase the risk in the same amount. Furthermore, the majority of breast cancers are not linked to mutations in these two genes. In other words, having the mutations raises the risk, but not having them does not lower it.

So, let's get some numbers straight. According to the American Cancer Society about 15% of women diagnosed with breast cancer have a family member diagnosed with it. That leaves the majority of breast cancers unrelated to family history:

"About 85% of breast cancers occur in women who have no family history of breast cancer. These occur due to genetic mutations that happen as a result of the aging process and life in general, rather than inherited mutations."

It's a puzzle I've discussed before, the missing herediatbility. On the one hand we know genes play a large role in cancer and we spend all this research money into looking for genetic causes. Yet, the vast majority of cancers are non-hereditary.

While women with certain mutations in either the BRCA1 or BRCA2 genes have up to 80% (the exact chance varies depending on the type of mutation they carry) increased risk of developing breast cancer, only between 5% and 10% of breast cancers are linked to deleterious mutations in the BRCA1 or BRCA2 genes. So, yes, get tested. But chances are, your copy of BRCA1 and BRCA2 are fine.

So, what makes BRCA1 and bRCA2 so scary?

The American Cancer Society reports that approximately 60% of women with one of the harmful mutations in BRCA1 or BRCA2 develop breast cancer during their lifetime, versus the 12% of women in the general population. Remember, though: these genes are not the only ones playing a role in cancer. Things like epistasis with other loci in the genome can deeply affect such risks and, unfortunately, we still don't know enough to quantify them. High levels of IGF-1, the insulin-like growth factor have also been linked to breast cancer. So while having those mutations raises the risk, it does not mean that the individual will develop breast cancer for sure as other factors are still unknown. Careful considerations should be made before making a drastic choice like Angelina's. These considerations should also include risks associated to a double mastectomy (infection, necrosis, etc.) and reconstruction surgery, neither one free of complications. I'm somehow reluctant to consider implants healthier than normal breasts, whether or not those breasts were expressing faulty genes.

What about those 85% of breast cancers that are not linked to BRCA1 or BRCA2 mutations? Can we do anything to prevent those?

When you look at the global population, the most common risk factors for breast cancer are not the mutations in BRCA1 and BRCA2, rather, as Bernstein reports in a 2009 review [2]:

"The most consistently acknowledged risk factors for breast cancer other than gender and race/ethnicity are age, family history of breast cancer, early menarche, late age at first birth, nulliparity, late age at menopause, high postmenopausal weight or substantial weight gain as an adult, exposure to high levels of ionizing radiation and a history of benign proliferative breast disease [2]."

All these risk factors point at one common etiology, ovarian hormones (estradiol and progesterone), because they

"promote cellular proliferation in the breast, providing greater opportunity for the accumulation of random errors, which may lead to tumor development [2]."

Body weight and exercise can be linked to different levels of estradiol in the blood (high body weight is associated with higher levels, exercise is associated with lower levels), hence their correlation to breast cancer risk. Some studies found up to 40% reduction in risk in women who exercised in particular in their adolescence. Of all risk factors, these two, body weight and exercise, are the ones we can actually take control over and actively lower our risk of developing breast cancer. A diet rich in antioxidants may lower the risk of DNA damage during cellular division.

Things we have less control over is the woman's age at the first pregnancy. One of my grad school professors used to say, "Having a baby as a teen may ruin your life, but it sure lowers your risk of developing breast cancer later in life." The risk keeps lowering for every additional pregnancy, though not as significantly as with the first one.

What's not clear is the extent to which breastfeeding can lower the risk of breast cancer, as the American Cancer Society reports:

"Research suggests that breastfeeding has only a slight effect on breast cancer risk and that effect is only among women who have breastfed for a long time. They also concluded that breastfeeding seems to be more protective against the most aggressive types of breast cancer, including tumors in women with mutations in the BRCA1 gene, basal-like cancers, hormone-receptor negative, and possibly triple negative tumors."

And while we do the things that we can to lower our risks, I am hopeful that one day gene therapy will be perfected to the point that it will offer a better options than what, in gross terms, amounts to amputation.

Thoughts?

[1] Hall, J., Lee, M., Newman, B., Morrow, J., Anderson, L., Huey, B., & King, M. (1990). Linkage of early-onset familial breast cancer to chromosome 17q21 Science, 250 (4988), 1684-1689 DOI: 10.1126/science.2270482

[2] Bernstein, L. (2008). Identifying population-based approaches to lower breast cancer risk Oncogene, 27 DOI: 10.1038/onc.2009.348

Pumping fuel from bacteria

In my last post I discussed a bioengineered E. coli strain capable of producing an engine compatible biofuel. I hailed the finding as more efficient than ordinary biofuels because this technique has less environmental impact than biofuels from crops, for example, or cellulose, which instead use great amounts of water and forest land.

I did some more reading on the topic and found out that, surprise surprise, there are some costs in harvesting biofuels from bacteria as well, so my discussion was incomplete. However, there are good news at the horizon.

When I first read the Howard et al. paper, I imagined a petri dish of E. coli sitting in a slime of oil-like substance. I think I got confused with making yogurt. :-) In reality, the biofuel molecules are stored inside the cells (bacteria, in this case) and need to be taken out without harming the cells. Biofuel secretion strategies have been dubbed "milking." The difference, though, is that contrary to milk and cows, biofuels are generally toxic to the bacteria that produce them.

Several methods have been investigated to efficiently "milk" biofuel molecules out of bacteria without harming them. To understand these strategies, we need to learn a new concept: an efflux pump is a membrane transporter protein that carries a substance toxic to the cell outside the cell itself. These proteins remove all kinds of toxic substances, including antibiotics, for example, and they may be specific to one in particular, or carry a whole range.

In [1], Dunlop et al. discuss the use of efflux pumps in "milking" biofuels out of bacteria and reduce their toxicity to the cells:

"Many compounds being considered as candidates for advanced biofuels are toxic to microorganisms. This introduces an undesirable trade-off when engineering metabolic pathways for biofuel production because the engineered microbes must balance production against survival. Cellular export systems, such as efflux pumps, provide a direct mechanism for reducing biofuel toxicity."

The researchers first looked at the whole genome of E. coli to identify all genes encoding efflux pumps. They found 43 different pumps expressed in the E. coli genome, and tested them against a range of possible biofuels. Their strategy was as follows: the grew a culture of pooled bacteria with different subpopluations, each subpopulation expressing a different pump. In the absence of toxic biofuel-like substances, all subpopulations grew in equal proportions, and none had an advantage over the others. When a substance was introduced, the subpopulations with the most advantageous pumps with respect to that particular substance outgrew the rest.

This is what happened, for example, when they introduced geranyl acetate:

"When the pooled culture was grown in the presence of an inhibitory biofuel such as geranyl acetate, some efflux pumps conferred a distinct advantage. Although all strains started out with equal representation, after 38 h the population composition changed, with cells containing the advantageous pumps becoming an increasingly large proportion of the population. The efflux pumps that enhanced tolerance to geranyl acetate originated from a variety of hosts and include both known and previously uncharacterized pumps."

In their study, Dunlop et al. used a type of membrane transporters called "RND," which are made of big molecules and are only found in Gram-negative bacteria. In a more recent paper [2], Doshi et al. studied a broader set of pumps called ABC, ATP-binding cassette:

"Unlike RND proteins, transporters belonging to the ATP- binding cassette (ABC) protein family are widely found in all five kingdoms of life. They share a conserved structural architecture and specifically import or export a wide variety of molecules and ions across cellular membranes."

Doshi et al. tested whether this family of broadly specific pumps could efficiently mediate the secretion of four different biofuel molecules. Similarly to the Howard et al. paper, they used a bioengineered strain of E. coli and noticed that

"the secretion process was sustained for at least 6 days without the need to replenish the growth medium or culture. Thus, for the same quantity of biofuel produced conventionally, we have a dramatic reduction in biomass scale and significant gain in the ease of recovering the biofuel."

Though my understanding is that work still needs to be done to improve this technique and make it feasible for different types of biofuels, the fact that these transporters are spread across different species makes it potentially translatable to other organisms and therefore of broader use.

On a completely different note, can you guess what the macro picture is? :-)

[1] Dunlop, M., Dossani, Z., Szmidt, H., Chu, H., Lee, T., Keasling, J., Hadi, M., & Mukhopadhyay, A. (2011). Engineering microbial biofuel tolerance and export using efflux pumps Molecular Systems Biology, 7 DOI: 10.1038/msb.2011.21

[2] Doshi, R., Nguyen, T., & Chang, G. (2013). Transporter-mediated biofuel secretion Proceedings of the National Academy of Sciences DOI: 10.1073/pnas.1301358110

Fill the tank, please. With bacteria!

I apologize if you've already heard about this, but the paper is really cool and I couldn't resist discussing it here.

Escherichia coli, or E. coli for brevity, is a bacterium normally associated with "bad" things like food poisoning. Even though most strains are actually harmless, even the CDC has a page dedicated to E. coli outbreaks. Since it's part of our gut flora, the lower intestines in particular, it's usually not a good sign when E. coli is found in places like restaurants and cafeterias. (Yuck!)

What's less known to the public is that E. coli is one of the most studied bacteria and makes a great model for mutations, gene duplications, and horizontal gene transfer.

What's even less known is that this amazing bacterium has the potential to save our planet from further drilling. How? By producing fuel. Yes, you read that right: through a combination of gene modifications, researchers from the University of Exeter [1] induced "petroleum-replica hydrocarbons" production in E. coli. These hydrocarbons are structurally and chemically similar to fossil fuels.

In their paper, Howard et al. argue against the current biofuels because they bring additional costs in downstream processing and are not 100% compatible with the engines on the market.

"To overcome the end-user blend wall, it is essential to generate precise chemical replacements to fossil fuels through sustainable means.Retail transport fuels are composed primarily of hydro- carbons (n-alkanes) of various carbon chain lengths (Cn), branched hydrocarbons (iso-alkanes), and unsaturated hydrocarbons (n- alkenes). The ideal biofuels are therefore n-alkanes, iso-alkanes, and n-alkenes that are chemically and structurally identical to the fossil fuels they are designed to replace [1]."

Gasoline, diesel and jet fuels are made primarily of molecules called alkanes, or saturated hydrocarbons. Most people are familiar, or at least have heard of methane, the simplest alkane molecule. These molecules are naturally produced not just by bacteria, but also by plants and insects when they metabolize fatty acids. In 2010 Schirmer et al. described in a Science paper [2] an alkane biosynthesis pathway in cyanobacteria, commonly known as blue-green algae.

"The pathway consists of an acyl-acyl carrier protein reductase and an aldehyde decarbonylase, which together convert intermediates of fatty acid metabolism to alkanes and alkenes [2]."

Understanding how alkanes are produced and, in particular, which genes are involved in their production, was the first step. The second step was answering the question: can we tweak this pathway to produce alkanes that can replace our current fuels?

Seen under this light, the PNAS study published last March 15 [1] is a bioengineering success story. Howard et al. designed a novel metabolic pathway that forced E. coli to use free fatty acids instead of fatty acid compounds as in cyanobacteria, and produce fuel-like alkanes, what the authors call "industrially relevant, petroleum replica fuel molecules." Once finalized, this type of biofuel will be compatible with current engines and will not need to be blended with other petroleum derived chemicals.

A bit of perspective: though derived from natural and biological sources, biofuels still contribute to pollution, carbon emissions, and global warming. Despite the amicable "bio" prefix, they all come with a non-null carbon footprint, some more than others. The true efficiency of any kind of fuel is the energy they produce minus the energy and costs it takes to derive them. For example, producing biofuels from crops drains precious resources, first and foremost, water, but also arable land, forests when arable land is not available, and food sources in underdeveloped countries.

So here's where biofuels from bacteria have a striking advantage: E. coli is one of the cheapest and easiest bacterium to grow in a lab. It doesn't drain water reservoirs and it doesn't need deforestation to grow. Contrary to most biofuels out there, that have high production and energy costs, the carbon footprint of biofuels derived from bacteria only comes from carbon emissions when you burn them.

And while this is an excellent thing, I still think that the real change we need to make to preserve our planet is to switch to renewable energy.


[1] Howard, T., Middelhaufe, S., Moore, K., Edner, C., Kolak, D., Taylor, G., Parker, D., Lee, R., Smirnoff, N., Aves, S., & Love, J. (2013). Synthesis of customized petroleum-replica fuel molecules by targeted modification of free fatty acid pools in Escherichia coli Proceedings of the National Academy of Sciences DOI: 10.1073/pnas.1215966110

[2] Schirmer, A., Rude, M., Li, X., Popova, E., & del Cardayre, S. (2010). Microbial Biosynthesis of Alkanes Science, 329 (5991), 559-562 DOI: 10.1126/science.1187936

And finally... spring! (And a plea for the bees)

A few days ago I complained about spring being late this year, so I thought I'd follow up with an update. :-)

Yup. I think it's here now. :-)

Sadly, I didn't see as many bees as I'd wished I did, and I fear it may not be a coincidence. Last year, my friend and colleague Bette Korber wrote a beautiful post on her blog and how we should all do our part to preserve these precious tireless workers:

"Bees are in trouble, and wild bees are disappearing. It's possibly caused by a lethal combination of virus and fungus, thought better able to take hold when bees are stressed and weakened. And stress them we do, with our pesticides, agricultural monoculture, and habitat loss. We can help the bees in a simple joyful way by planting native wildflowers in our gardens, different kinds, in abundance. Pesticide-free bee habitat restoration, one yard at a time. Spring is coming – get busy."

North Korea and the USA can indeed unite: in the battle against TB.

Tuberculosis (TB) is an infectious disease caused by a bacterium. It spreads through cough or sneeze from subjects with an active infection. While in most cases the disease is asymptomatic, a minority of latent infections does become active (i.e. the subject develops symptoms), and when it does, if left untreated, the disease can be deadly.

According to the CDC one third of the world's population are infected with TB, and while in the US the incidence of the disease has been declining over time, it is still a huge problem in parts of the world like Asia and sub-saharan Africa. While normally the chance of a latent TB infection becoming active is one in ten, the chance is much higher for HIV-positive subjects because their immune system is already debilitated by the HIV virus. As the CDC reports:

"TB is a leading killer of people living with HIV (PLHIV)."

A regimen of 3-4 drugs has been available for years to keep latent infections from becoming active. Sadly, TB infections from multidrug resistant strains (MDR) have been steadily increasing, setting back the progress made in the past decades.

From the World Health Organization:

"Drug resistance arises due to improper use of antibiotics in chemotherapy of drug-susceptible TB patients. This improper use is a result of a number of actions including, administration of improper treatment regimens and failure to ensure that patients complete the whole course of treatment. Essentially, drug resistance arises in areas with weak TB control programmes. A patient who develops active disease with a drug-resistant TB strain can transmit this form of TB to other individuals."

One of the countries plagued by MDR TB strains is North Korea, where the incidence of TB has dramatically advanced over the past years, reaching one of the highest incidences outside sub-saharan Africa.

In this week's issue, Science Magazine describes a joint effort between two countries that, according to the recent news, you'd least expect to pair up: North Korea and the United States. In collaboration with Stanford University, the Korean ministry of Public Health opened in 2010 a National Tuberculosis Reference Laboratory (NTRL).

"NTRL researchers can now diagnose TB cases that are resistant to first-line drug combinations, making it possible to spot patients who need more aggressive therapy. And the lab will soon add capacity to screen for extensively drug-resistant TB, known as XDR—the worst strains, some of which are close to impossible to treat."

The Science report covers stories of hope in the midst of desperation. It points to pressing issues the North Korean government has to address within its borders, and focusing on them would seem a more reasonable and logical strategy than polishing nuclear arsenals. Let's hope that the roots of this collaboration grow deeper than any political discrepancies. Let's hope that the battle against a common enemy (TB) will put an end to the empty, unfounded threats and pave the way to a broader, more civilized way of communication between countries.

Stone, R. (2013). Public Enemy Number One Science, 340 (6131), 422-425 DOI: 10.1126/science.340.6131.422

Who is Publishing in My Domain?

Work mornings always start with the same task: sort through email. Lately, I've been getting a lot of uncategorized junk, mostly invitations to conferences that have nothing to do with my work (though I'd love to go to China, Thailand, Abu Dhabi, etc., I typically need a valid justification to ensure my employer pays for it), and labs who haven't figured out that I'm a theoretician, not an experimentalist, and send me special offers on antibody assays, genetic typing, ELISA, etc.

Needless to say, my index finger has developed a "junk mail" reflex that spares me to even open such emails.

So this morning I almost deleted an email that actually put a smile on my face. Small paper, small accomplishment, but in a challenging job, it's the small things that keep one going:

===================================
BioMedLib: "Who is Publishing in My Domain?"
===================================

The following section is the top 20 articles published on the same topic since you published yours.

List 1: Top 20 Articles, in the Domain of Article 20973976, Since its Publication (2010)

1. Estimating time since infection in early homogeneous HIV-1 samples using a poisson model.
Giorgi EE, Funkhouser B, Athreya G, Perelson AS, Korber BT, Bhattacharya T.
BMC Bioinformatics; 2010;11:532.

2. Statistical inference of selection and divergence from a time-dependent Poisson random field model.
Amei A, Sawyer S.
PLoS One; 2012;7(4):e34413.

3. A model of HIV-1 infection with two time delays: mathematical analysis and comparison with patient data.
Pawelek KA, Liu S, Pahlevani F, Rong L.
Math Biosci; 2012 Jan;235(1):98-109.

4. ...

Yup. Ours was the first in the list. Doesn't happen every day, does it? In fact, I think I'm gonna cherish this moment since I'm pretty sure it won't happen again. :-)

PS Now, don't you go around telling me that there's no value in such stuff, I know there isn't, but one's gotta believe in something, right ??

Can we functionally cure HIV?

Last March, Dr. Deborah Persaud, from the John's Hopkins Children Center, presented a stunning finding at the conference CROI, receiving great resonance across several newscasts: Persaud reported the first case of infant functionally cured of HIV. You can watch Persaud's presentation by downloading the podcast here, it's the seventh talk of the session "Is there hope for HIV eradication?"

Up until this finding, the only living person cured from HIV was the Berlin Patient, who was cured after receiving gene therapy for his underlying leukemia condition. Despite this one successful case, gene therapy is not a feasible way to cure HIV.

What does it mean to be functionally cured?

Once in the host, the HIV virus establishes reservoirs of latent virus: these are viral particles that stay dormant in cells and tissues and have the ability to quickly rebound in the event that therapy is discontinued. That's why it's so important for an HIV infected person to never discontinue the drug regimen, as the rebound virus may be drug resistant. HIV is so efficient at escaping the immune system and therapy that standard practice these days is a lifetime of not just one, but a cocktail of 3-4 antiretroviral drugs.

To be functionally cured means that drugs are no longer needed to keep the viral load (amount of virus in the blood) in check (close or below detection), something that until now had only been achieved by an extremely low number of HIV-positive individuals (less than 1% of infected adults), the so-called "elite controllers." In all other subjects, the reservoirs are never completely weakened and they enable the virus to bounce back once therapy is interrupted.

So, what was different with this child?

The mother went into labor without prenatal care. An HIV test was done during labor and normally, when the test is positive, antiretroviral drugs are administered. This is highly effective in preventing mother-to-infant infections as the only moment when the infant is exposed to the mother's blood is at birth. The antiretroviral drugs keep the viral load so low that the risk of infection becomes very small (around 2%). Unfortunately, in this particular case, the birth was so precipitous that there was no time to administer such drugs. The newborn baby was immediately tested for HIV.

This is my understanding of what was unique about this case: normally a first test is done and, if positive, a second follow-up test is performed and prophylaxis is started once the infection is confirmed. In this case, though, two independent tests were done at the same time and, since both confirmed the HIV infection, prophylactic treatment was started very early, when the baby was 31 hours of age. Also, unique to this case was the fact that a regimen of three drugs, of which one at the therapeutic level instead of the standard prophylactic dosage, was administered during the first week of life. After that, the baby was switched to a standard treatment of antiretroviral drugs (again, my understanding from the CROI talk).

Such regimen successfully brought the child's viral load down to undetectable, which is normal in these cases. Despite this, because of HIV's ability to establish reservoirs, antiretroviral therapy is never discontinued. Like I said before, it is a lifetime therapy. So called "drug holidays" result in more virulent and drug-resistant HIV quasispecies. However, this child was lost to follow-up at 18 months of age and was once again seen by the doctors at 25 months of age, when the caregiver reported discontinuing the therapy. Immediate testing was done to assess the child's viral loads. The child was tested not once, but many times. Genetic testing was also done to make sure it was the same child treated before. The doctors must have been in disbelief as for the first time they were seeing the incredible: after 5 months since discontinuing antiretroviral therapy, the viral load in this child was still undetectable.

What are the consequences? As Dr. Persaud repeated many times during her talk, this is a single case and a proof of concept. We need more cases to be able to generalize (as statistics teach us). However, it points to something that indeed needs to be explored: how early in the infection can we (and should we) intervene? In a 2012 paper [1], Persaud and colleagues studied the dynamics of the latent HIV reservoirs in 17 infants on very early antiretroviral drug therapy (median start age 8 weeks) and found that the size of the reservoirs at age 2 was associated to how early undetectable viral loads were achieved during therapy. The earlier viral load was suppressed through therapy, the smaller the HIV reservoir at age 2. Is there a point, very early into the infection, when the virus is vulnerable and all reservoirs can be not just reduced in size, but actually completely eradicated through potent and prompt intervention?

In rare cases, HIV-infected patients are able to spontaneously maintain their viral load at a very low level without the need of drugs, the so called "elite controllers." What if, when administered early enough, antiretroviral drugs could transfer this type of spontaneous protection to every HIV-infected person?

Shortly after the CROI conference, a French study published in PLoS Pathogens [2] reported 14 cases of what they call "post-treatment controllers," in other words, people whose viral loads remained very low after interrupting treatment. With the exception of mother-to-infant transmissions at birth, it's extremely hard to catch this virus early because people often don't realize they've been infected: symptoms, if any, appear 3-4 weeks later and are often mistaken for a common cold. Twelve of the 14 cases reported in [2] had symptoms that prompted early intervention and start of therapy during the primary infection.

"Post-treatment controllers (PTCs) had a more severe primary infection with higher viral loads and were frequently symptomatic, which may have prompted the early treatment in some cases [. . .] Therefore, our results strongly suggest that the infection control in the PTCs was not achieved spontaneously and was favored by the early onset of therapy. Because the interruption of long-term antiretroviral therapy initiated early during primary infection is not recommended, only a very small proportion (~2%) of the patients in the French Hospital Database on HIV Infection experienced such an interruption, which may explain the rarity of PTCs worldwide [2]."

[1] Persaud, D., Palumbo, P., Ziemniak, C., Hughes, M., Alvero, C., Luzuriaga, K., Yogev, R., Capparelli, E., & Chadwick, E. (2012). Dynamics of the resting CD4+ T-cell latent HIV reservoir in infants initiating HAART less than 6 months of age AIDS, 26 (12), 1483-1490 DOI: 10.1097/QAD.0b013e3283553638

[2] Sáez-Cirión, A., Bacchus, C., Hocqueloux, L., Avettand-Fenoel, V., Girault, I., Lecuroux, C., Potard, V., Versmisse, P., Melard, A., Prazuck, T., Descours, B., Guergnon, J., Viard, J., Boufassa, F., Lambotte, O., Goujard, C., Meyer, L., Costagliola, D., Venet, A., Pancino, G., Autran, B., Rouzioux, C., & , . (2013). Post-Treatment HIV-1 Controllers with a Long-Term Virological Remission after the Interruption of Early Initiated Antiretroviral Therapy ANRS VISCONTI Study PLoS Pathogens, 9 (3) DOI: 10.1371/journal.ppat.1003211

Waiting for spring...

No sign of spring here in Northern New Mexico, so I brought in some flowers from Southern California. I didn't have the macro with me, these are all shot at 200mm, two stops down to make the background dark.

Science is sexy. And to prove it, we show you 50 sexy scientists.

No, I'm not joking. It's real, check it out:

50 Sexy Scientists

So, what do you think? Would you be flattered to be on that list?

I find it a little disturbing. Giving the scientist's looks the same merits as his/her work is like comparing pears with oranges. Or, like comparing liters with watts. In fact, I think it promotes a disturbingly negative message, which is: do I need to be beautiful to have my work noticed?

What do you think? Am I over-reacting?

NOTE: Link found via A.V. Flox.

Antiviral drugs to fight the flu: yes or no?

Disclaimer: I'm not a medical doctor. I cannot recommend taking or not taking a certain drug. However, I am a human being, I've got kids who do get sick from time to time, and I work on viruses. So when I heard that people were battling the unusually nasty flu this year with antiviral drugs, well, I had to do a bit of research.

Antiviral drugs have become increasingly popular after the highly pathogenic avian flu strain emerged. The idea is that in order to be prepared for a possible pandemic, we need to stock up on drugs, enough to treat millions of people.

Let's start with a few facts about viral infections:

A virus is made of genetic material packaged in a tiny shell. Once inside a cell, the virus hijacks the cell's proteins to replicate and create thousands of copies of itself. The new virions bud out of the cell membrane and infect new cells.

However, the infected cell has one more weapon up its sleeve: it "grasps" the virions that are budding out and tries to hold them back. In more scientific terms: there's a molecule on the cell membrane (called sialic acid) that binds to a protein on the surface of the virus (called hemagglutinin). The new virions have to break this bond in order to leave the infected cell and spread the infection. How do they do that? They use an enzyme called neuraminidase: the enzyme, found on the surface of the virus, breaks the bond between hemagglutinin and siliac acid, setting the virions free to spread out and infect new cells.

Neuraminidase inhibitors are antiviral drugs that, as the name suggests, block the neuraminadase enzyme. As a consequence, the virions remain stuck to the cell membrane and thus cannot spread the infection. Supposedly, this leads to a speedier recovery. I'm saying "supposedly" because there is an ongoing debate on whether this is true or not. Here's what I found in the literature.

Jefferson et al., 2009 [1]:

"Neuraminidase inhibitors have modest effectiveness against the symptoms of influenza in otherwise healthy adults. The drugs are effective postexposure against laboratory confirmed influenza, but this is a small component of influenza-like illness, so for this outcome neuraminidase inhibitors are not effective. Neuraminidase inhibitors might be regarded as optional for reducing the symptoms of seasonal influenza. Paucity of good data has undermined previous findings for oseltamivir's prevention of complications from influenza. Independent randomised trials to resolve these uncertainties are needed."

Wang et al., 2012 [2]:

"Oseltamivir and zanamivir appear to have modest benefit in reducing duration of illness in children with influenza. However, our analysis was limited by small sample sizes and an inability to pool data from different studies. In addition, the inclusion of data from published trials only may have resulted in significant publication bias. [...] The clinical efficacy of neuraminidase inhibitors in 'at risk' children is still uncertain. Larger high-quality trials are needed with sufficient power to determine the efficacy of neuraminidase inhibitors in preventing serious complications of influenza (such as pneumonia or hospital admission), particularly in 'at risk' groups."

Jefferson et al. 2012 [3]:

"We found a high risk of publication and reporting biases in the trial programme of oseltamivir. Sub-population analyses of the influenza infected population in the oseltamivir trial programme are not possible because the two arms are non-comparable due to oseltamivir's apparent interference with antibody production. The evidence supports a direct oseltamivir mechanism of action on symptoms but we are unable to draw conclusions about its effect on complications or transmission. We expect full clinical study reports containing study protocol, reporting analysis plan, statistical analysis plan and individual patient data to clarify outstanding issues. These full clinical study reports are at present unavailable to us."

The company that manufactures the brand name drug for oseltamivir responded here, however, according to BMJ, there hasn't been any release of data yet (source).

I have two more cautionary comments.

First: viruses mutate very rapidly and as such, they rapidly find escapes to drugs. An overuse of antiviral drugs may end up selecting drug resistant strains (for example, a flu strain that carries a neuraminadase enzyme that the inhibitor drugs cannot block). I'm not saying that antiviral drugs should not be used. Some life-threatening situations require the use of such drugs (for example, in the case of patients with immunodepression). Other non life-threatening situations don't (plain and simple).

Second: the US Food and Drug Administration recommends the use of antiviral drugs and in fact, last December they expanded the recommendation to children under one year of age (source). However, if you keep browsing the FDA website you find this very interesting Q&A page, where they report some supposed (though worrisome) adverse side effects:

"In the safety review mandated by the BPCA, a number of adverse event reports were identified associated with the use of Tamiflu in children 16 years of age or younger. These adverse event reports were primarily related to unusual neurologic or psychiatric events such as delirium, hallucinations, confusion, abnormal behavior, convulsions, and encephalitis. These events were reported almost entirely in children from Japan who received Tamiflu according to Japanese treatment guidelines (very similar but not identical to U.S. treatment guidelines). The review identified a total of 12 deaths in pediatric patients since Tamiflu's approval. All of the pediatric deaths were reported in Japanese children. In many of these cases, a relationship to Tamiflu was difficult to assess because of the use of other medications, presence of other medical conditions, and/or lack of adequate detail in the reports."

There is no direct evidence that the deaths were linked to the use of the drug. In fact, often it's high risk children that need to take the drug, which means they are likely to have other conditions and take additional medications. If we can't be certain of what one drug alone can do, imagine multiple ones combined. You can read more about the Japan reports here. I also found a reference [4].

Bottom line: drugs are wonderful things. They save lives. Drugs can also mess up with our body chemistry in ways that we don't always understand. The key point is to read, be informed, and use sparingly (as needed, not just as recommended).

NOTE: I strive to make these commentaries as objective as possible. If you feel I've missed some part of the story or you have more references to add to make a rounder point, please let me know in the comments. 

[1] Jefferson, T., Jones, M., Doshi, P., & Del Mar, C. (2009). Neuraminidase inhibitors for preventing and treating influenza in healthy adults: systematic review and meta-analysis BMJ, 339 (dec07 2) DOI: 10.1136/bmj.b5106

[2] Kay Wang, Matthew Shun-Shin, Peter Gill, Rafael Perera, Anthony Harnden (2012). Neuraminidase inhibitors for preventing and treating influenza in children (published trials only) The Cochrane Library DOI: 10.1002/14651858.CD002744.pub4

[3] Tom Jefferson, Mark A Jones, Peter Doshi, Chris B Del Mar, Carl J Heneghan, Rokuro Hama, Matthew J Thompson (2012). Neuraminidase inhibitors for preventing and treating influenza in healthy adults and children The Cochrane Library DOI: 10.1002/14651858.CD008965.pub3

[4] Urushihara, H., Doi, Y., Arai, M., Matsunaga, T., Fujii, Y., Iino, N., Kawamura, T., & Kawakami, K. (2011). Oseltamivir Prescription and Regulatory Actions Vis-à-Vis Abnormal Behavior Risk in Japan: Drug Utilization Study Using a Nationwide Pharmacy Database PLoS ONE, 6 (12) DOI: 10.1371/journal.pone.0028483

San Francisco Bay Bridge

I recently got back from a 5-day trip to the Bay Area, where, over the week-end, I was privileged to go shooting with a group of local photographers. Since I don't have much water here in NM, the thing I enjoyed the most was doing long exposures by the water.

I wanted to show you some examples of shots I took of the Bay Bridge. I do some editing, mainly I add contrast, lower highlights and tweak exposure as needed, but what makes these shots so different is the timing. As the light changes, so do the colors. I did use a graduated neutral density filter, which does not change the colors. All it does is darken the upper portion of the frame so that you can achieve a more uniform exposure (otherwise the sky would be too bright).

For me, the lesson learned here is that the best time to shoot is blue hour, right after sunset. And you have to be quick because the colors change by the minute. I've noted the time each shot was taken below. Which one do you like best?

Right after sunset, 5:32 pm:

5:33 pm:

5:39 pm:

5:47 pm:

5:54 pm:

Different day, much later: 6:44 pm. As you can see, the pastel colors are pretty much gone and all is left is yellow and blue-purple. I tried to fiddle with white balance, but got stuck with pretty much those colors. No filters here as it was already quite dark.

Much later at night, 12:56 am (in this one I switched to tungsten to drop the color temperature):

1 am (standard white balance, this time, but I tweaked temperature and tint):

I'm self-taught, so I'm learning all this stuff by myself (well, not quite, I'm learning from other photographers!). I welcome critique/tips/advice for next time. Thanks!