Monthly Archives: October 2014

Genome Browser in a Box (GBiB) Origins

The opinions expressed here are those of the author, Jonathan Casper, and do not necessarily reflect those of the University of California Santa Cruz or any of its units.

I’m happy to say that we’ve finally released the Genome Browser in a Box (GBiB). GBiB is essentially a virtual machine image of a mirror of the UCSC Genome Browser. Download it, set it up, and voilà – instant mirror. This lets you do cool things like have a mirror of the browser on your own personal computer – more information on how to set it up is available on the help page at http://genome.ucsc.edu/goldenPath/help/gbib.html. Here, however, I’m going to talk about the background of GBiB: how it got started, what kinds of decisions we faced, and what became my favorite feature.

At UCSC, we have known for a long time that it can be hard to set up a mirror server. We even have a separate mailing list devoted to the topic. Before the GBiB project began, one of our developers was working on a script to completely automate the process for a computer running stock Ubuntu Linux.  Eventually the developer had an epiphany: why not just create a barebones mirror once on a virtual machine, and then make that available to our users? It wouldn’t solve the problem of allowing people to easily add the assemblies and tracks they wanted, but at least they wouldn’t have to wrestle first with setting up Apache and MySql. The developer’s suggestion came at an opportune moment – we had just received several mailing list questions about using sensitive data with the UCSC Genome Browser website. We didn’t have a good answer.

The problem is that the UCSC Genome Browser has always been focused on being an academic research tool, not a clinical one. We aren’t designed to provide the kind of data security that HIPAA and Institutional Review Boards call for. The only answer we could give to people who wanted data security was “create your own secure mirror, or use another genome browser”. Knowing how difficult it could be to set up a mirror, that wasn’t much of a choice.

Into that mix, we were suddenly presented with a new option: give everyone a pre-installed mirror with the hardest parts already done. Just place it behind a firewall, load up your sensitive data, and enjoy! I thought it was a great idea, as did many other browser staff members.

From there, the idea quickly snowballed. UCSC already provides a public MySQL server and download site with most of the data from our browser. We realized that we could set up the virtual machine to take advantage of those resources and load our data over the internet. This was a great advantage over normal mirror servers. UCSC provides many terabytes of data. Most mirrors have to pick and choose which assemblies and tracks they make available; there’s far too much data to download and keep synchronized. By using our public internet resources, GBiB could provide all of it.

In practice, we discovered it wasn’t quite that easy. Latency issues meant that for anyone not on the west coast of the United States, GBiB worked really slow. Just loading the default view of the human GRCh37/hg19 genome assembly could take over 10 seconds. We had to make a compromise: GBiB wouldn’t have to download track data to use it, but downloading would still be an available option for users in remote locations.

There is now a new CGI just for this purpose: “Mirror Tracks”. It combs through the list of database tables and files associated with browser tracks and allows you to download the data for any of them. If you’re interested in looking at, say, mRNA alignments in the Painted turtle (chrPic1) genome and GBiB is just too slow, a few clicks in Mirror Tracks will put them all on your own hard drive. If you really want, you can even then put GBiB into full-offline mode. You’ll lose access to any track data that you haven’t downloaded, but you’ll always have those Painted turtle mRNAs.

My favorite feature of GBiB, though, has to be what it does for track hubs. Track hubs are a feature we released in 2011 to allow users to view their own data files in the UCSC Genome Browser alongside our annotation. Unlike custom tracks, where all the data must be sent to our server at once, track hubs only send the data for the region you are looking at. That is much more manageable for something like a VCF file, which can be on the order of 10-100 GB.

There are two problems with track hubs. First, you must have web hosting space for your data to construct a track hub. Not everyone does. There are public hosting solutions like DropBox, but they don’t always work. Second, once again there is the problem of sensitive data. Even if you are willing to send your sensitive data directly to our servers at UCSC, you may not be willing (or even allowed) to make it publicly available on a web server. GBiB solves both problems beautifully.

GBiB already has a built-in web server that it uses to communicate with your computer. With a few small adjustments, you can take advantage of that and let GBiB also host your data files. This means that you can build and use a track hub with GBiB, and none of the data will ever leave your computer or be accessible to anyone else, unless you grant them access to your GBiB.

Genome Browser in a Box is available from our web store at https://genome-store.ucsc.edu. It is free for non-commercial use by non-profit organizations, academic institutions, and for personal use. Please see the store website for full terms and conditions.

2014 Ebola epidemic

The opinions expressed here are those of the author, Jim Kent,  and do not necessarily reflect those of the University of California Santa Cruz or any of its units.

I first learned about Ebola in a microbiology course in 1997. This was a great course and the teacher was very funny. He joked that our mothers taught us to wash our hands after we went to the bathroom, but in this class it was a good idea to wash hands before going to the bathroom. He once held up a test tube of Clostridium botulinum and joked it was enough to wipe out a day care center. He didn’t make any jokes about Ebola.

Ebola has been around in the African rain forests for a long time. There is some animal, probably a fruit bat, but nobody is really sure, that is the primary host of this virus. The disease a virus causes in its primary host is usually pretty mild, since when the host dies, the virus dies too. The virus can infect other animals as well. Dogs in the regions often have antibodies indicating exposure, but apparently it does not make them very sick at all, and they clear the actual virus from their systems quickly. Pigs do get sick, but generally recover. For chimps, gorillas, and humans it is usually fatal, particularly the Zaire strain of Ebola that is currently ravaging West Africa.

In humans Ebola is a disease that moves fast. It tends to either kill you quickly, or you survive it and clear it from your system relatively quickly. For a long time Ebola outbreaks affected only humans in remote villages in the forest, the ones most likely to come in contact with infected animals in the wild. The disease would burn through the village, and by the time survivors, if there were any, managed to come into contact with other people, they were non-infectious.

As Africa grew more populated and villages turned into towns, Ebola became a greater problem. People infected with Ebola and still in a contagious state would manage to stagger into a medical facility seeking care. The Nova episode Ebola, the Plague Fighters documents one such case. The patient arrived in 1995 in a hospital in Kikwit, Zaire (now DRC) with extreme pain in the abdomen. The doctors suspected appendicitis and operated. In spite of the usual surgical care against infection, nearly all of the medical personnel at that operation got infected and spread it throughout the hospital. It took heroic efforts to contain the resulting outbreak, which resulted in 315 infections and 244 fatalities.

There have been 7 Ebola outbreaks infecting 100 or more people since Kikwit. With moats of bleach-water, aggressive measures to trace contacts and quarantine people infected, and with the growing experience and energetic activities of groups such as the Doctors without Borders, each of these outbreaks has been contained. These outbreaks have, fortunately, provoked enough concern in the research community that vaccine and drug development is far along.

The 2014 outbreak in West Africa started in Guinea, and initially it looked like it would be contained as well. From what we can reconstruct, this one started with a two-year-old in December of 2013. The two-year-old survived, but the virus infected his family and many other people. The Guinea local health officials noticed the problem before it was too large and called in Médecins sans Frontières, as the Doctors without Borders are known in French (and indeed they prefer to use the French initials internationally, MSF). The MSF responded promptly and, working with locals, were able to contain the outbreak to just over 100 infected. Baize et al., 2014, in the New England Journal of Medicine, describe the course of this outbreak well and include sequence from 3 viral genomes from this outbreak.

For more than 42 days, long enough for two of the usual quarantine periods for Ebola to pass, there were no more cases. Then, in approximately the same places, the disease resurged. The reason for this resurgence is unknown. Sequencing of the resurgent outbreak by Gire et al., 2014, makes it clear that the resurgence is a continuation of the previous outbreak. It is possible that in this recently war-torn region, near the borders of Guinea, Sierra Leone, and Liberia, some of the infected fled and hid where MSF and the Guinea health workers never even knew about them. It is possible that local wildlife, perhaps a ground-based secondary host, acted as a reservoir for the virus.

Regardless of the cause of the resurgence, it has happened, and it has grown large, to the point that this is not an outbreak, but an epidemic. Ebola has for the first time hit densely populated regions. The epidemic has grown large enough that for all of their dedication and talent, MSF and similar organizations simply don’t have enough doctors and other health workers to contain it. The doubling time inside the worst hit city, Monrovia, Liberia, is just 2 weeks. The CDC’s best estimate is that 21,000 people were infected as of September 30. There is a very real danger that this epidemic could spread throughout Africa. There is a possibility that can’t be discounted, that in spite of the better sewers and other sanitation systems, it could spread through the developed world as well.

There are two things that are necessary to avoid a global pandemic. First, aggressive quarantine and isolation measures must be taken to slow the spread. Second, vaccines and treatments developed in response to previous outbreaks must be quickly scaled up so that hundreds of thousands, and ideally millions of doses are available. If either of these two things fails, we will face a worldwide problem of a scope that has not been seen for generations.

Amidst this gloom and doom there is some hope. Vaccine developers have already pressed two vaccines forward as far as they could go without actually having a human epidemic to test against. They work well in non-human primate trials. Comparative genomics analysis of the virus over the course of many outbreaks and across many strains makes it clear that there are large, antibody-accessible, parts of the virus that are highly constrained evolutionarily, and that the virus mutates slowly compared to HIV. This is something easy to see in the UCSC Genome Browser (http://genome.ucsc.edu/ebolaPortal/). It is exceedingly likely from the slow rate of change that both vaccines will work, and that once they are manufactured at large scale it will become much easier to contain the epidemic.

Similar logic applies to the treatment options that are under development. The ZMapp antibody cocktail tested in non-human primates and found effective against earlier versions of this virus likely will work in humans against the current strain. Likely, but not certainly. The Tekmira liposome-encapsulated siRNA is also a hopeful option, and perhaps can be scaled up faster than ZMapp, and perhaps also will work in humans. There is also hope that an existing drug, Lamivudine, a nucleoside analog currently approved for use against HIV and Hepatitis B, will be effective. Dr. Gorbee Logan in Liberia has had success using it on Ebola patients, and NIAID is following up on this with investigation in the lab. I’m hopeful the Ebola Genome Browser that UCSC just released is helpful for others developing new treatments.

Still, without effective quarantine measures this epidemic will grow very large indeed before vaccines and treatments can be deployed. The CDC estimate is 1.4 million cases by January 20, 2015, in Liberia and Sierra Leone alone, unless the rate of infection is slowed. Obama has ordered the army to deploy, and this is likely to help. From what I understand the plan is for the army to build field hospitals, and to help distribute food, water, and other necessary things, including vaccines and medicines once they are available.

Nonetheless I remain deeply concerned. It doesn’t really seem like people realize just how contagious this disease is. There’s thinking that somehow what is happening in Africa won’t happen elsewhere, that the African-specific customs and lifestyles favor the spread. While there is a grain of truth in this, it’s only a grain. We are fortunate in the developed world to have much better sewage systems than in Africa. We tend to leave it to professionals to prepare a body for burial rather than washing it ourselves. However, you only need look at how fast a cold or the flu passes around in the USA to see that we are not immune to a quickly progressing epidemic.

There have been many statements by the CDC and others, based on rather thin scientific evidence, that Ebola spreads only by direct contact with bodily fluids.   From this people seem to get the notion that if you avoid touching pools of blood, diarrhea, or vomit, you are ok.   Please let me take the opportunity to correct this notion.

The scientific evidence such as it is shows that if one cage of monkeys is infected with Ebola by injecting the virus into muscle tissue, it won’t infect a nearby cage in the same room in a set-up made to minimize large droplet transmission. In contrast with the same set-up, oro-nasally infected pigs were able to infect a nearby cage of monkeys in the same room (Weingartl et al., 2012). Earlier studies with a more casual layout (Jaax et al., 1995) did show indirect transmission between non-human-primates. For me, at least, the question of whether Ebola can infect without direct touch or large droplet contact remains unanswered.  At the least I would like to see if the monkeys infected by the pigs ora-nasally could in turn pass on the virus to other monkeys in the Weingartl et al. set-up. The site of initial infection has a profound effect on the progress and mode of transmission of many diseases.

Beyond this, even for the cold and the flu, 70% of infections are via contact. See the Influenza B article by La Rosa et al., 2013, and the review by Cowling et al., 2014, for further information on the modes of transmission of common viruses. What’s more, Ebola is a robust virus. It survives well outside of the body, particularly indoors away from the sun, compared to many common viruses (Piercy et al., 2010). You really need only touch something that someone with Ebola has touched recently. Among the bodily fluids that contain the virus is sweat!

Fortunately, there is no evidence that people are infectious before symptoms show. In macaques, sensitive RT-PCR tests are unable to detect the virus outside of the site of infection until day three post-infection, the same day symptoms develop, and even on day three, the levels are low, 1/10,000 of the levels they reach by day six (Giesbert et al., 2003). In humans we don’t have presymptomatic RT-PCR data, but we see similar patterns of a massive increase in viral load from the first symptomatic day to later in the disease (Towner et al., 2004). In all, the evidence that people are not infectious before showing symptoms seems solid.

Still, in the light of the relative ease of transmission of this virus once symptoms show, and the deadliness of the virus once caught, I simply don’t understand why travelers from the Ebola-stricken nations are not being quarantined. A single air traveler from Liberia to Lagos managed to infect 11 people directly, and 8 more indirectly. The Nigerian CDC was able to contain the outbreak, but it involved the efforts of hundreds of people tracing 900 contacts, and in the end 8 people died. A quarantine is a small price to pay to avoid situations like this. Recently a traveler from Liberia has developed symptoms in Dallas, and 100 contacts are being traced. We can only hope no fatalities will result.

It is my hope that people will start giving the virus the respect it deserves. I hope that all flights out of the infected regions will cease other than military flights where decontamination of planes and quarantine of passengers can be insured. I hope that sensible public health measures such as these will give us time to deploy vaccines and treatments, and develop backup vaccines and treatments in case the first set don’t work.

Meanwhile, back in West Africa, in Monrovia in particular, the epidemic has gotten so far that local quarantine is not sufficient. It has reached the stage where we have to start isolating the healthy into refuges. There are not enough health workers to handle the situation now, and the situation will rapidly get worse. The best we can do is to help people take care of their loved ones safely – distributing kits that ideally would include adult diapers, vomit buckets, gloves and breathing masks to help contain the spread of infected fluids; food, water, and electrolytes to support the patient; chlorine to disinfect the area and — if the Lamivudine treatment does hold up to its initial promise — a 5-day treatment of Lamivudine for the Ebola and a 5-day treatment of Amoxicillin to help contain the secondary infections. Kits such as these could end up saving hundreds of thousands of lives, especially if they can be delivered by people who know the local language and customs.

Many people in the world are doing all they can to fight this epidemic. It is inspiring to see. Through our combined efforts, I’m sure that the worst-case scenario won’t happen. Nonetheless what will happen is likely to be pretty bad if vaccine productions lag or if people don’t become a bit more realistic about just how infectious this disease is.

 

References:

Baize S, Pannetier D, Oestereich L, Rieger T, Koivogui L, Magassouba N, Soropogui B, Sow MS, Keïta S, De Clerck H et al. Emergence of Zaire Ebola Virus Disease in Guinea. N Engl J Med. 2014 Apr 16. doi: 10.1056/NEJMoa1404505

Cowling BJ, Ip DK, Fang VJ, Suntarattiwong P, Olsen SJ, Levy J, Uyeki TM, Leung GM, Peiris JS, Chotpitayasunondh T, Nishiura H, Simmerman JM. Modes of transmission of influenza B virus in households. PLoS One. 2014 Sep 30;9(9):e108850. doi: 10.1371/journal.pone.0108850. eCollection 2014.

Geisbert TW, Hensley LE, Larsen T, Young HA, Reed DS, Geisbert JB, Scott DP, Kagan E, Jahrling PB, Davis KJ. Pathogenesis of Ebola hemorrhagic fever in cynomolgus macaques: evidence that dendritic cells are early and sustained targets of infection. Am J Pathol. 2003 Dec;163(6):2347-70.

Gire, SK, Goba, A, Andersen, KG, … Sabeti PC et al. Genomic surveillance elucidates Ebola virus origin and transmission during the 2014 outbreak. Science 345, 1369–1372. doi:10.1126/science.1259657

Jaax N, Jahrling P, Geisbert T, Geisbert J, Steele K, McKee K, Nagley D, Johnson E, Jaax G, Peters C. Transmission of Ebola virus (Zaire strain) to uninfected control monkeys in a biocontainment laboratory. Lancet. 1995 Dec 23-30;346(8991-8992):1669-71.

La Rosa G, Fratini M, Della Libera S, Iaconelli M, Muscillo M. Viral infections acquired indoors through airborne, droplet or contact transmission. Ann Ist Super Sanita. 2013;49(2):124-32 .doi: 10.4415/ANN_13_02_03.

Piercy TJ, Smither SJ, Steward JA, Eastaugh L, Lever MS. The survival of filoviruses in liquids, on solid substrates and in a dynamic aerosol. J Appl Microbiol. 2010 Nov;109(5):1531-9.

Towner JS, Rollin PE, Bausch DG, Sanchez A, Crary SM, Vincent M, Lee WF, Spiropoulou CF, Ksiazek TG, Lukwiya M, Kaducu F, Downing R, Nichol ST. Rapid diagnosis of Ebola hemorrhagic fever by reverse transcription-PCR in an outbreak setting and assessment of patient viral load as a predictor of outcome. J Virol. 2004 Apr;78(8):4330-41.

Weingartl HM, Embury-Hyatt C, Nfon C, Leung A, Smith G, Kobinger G. Transmission of Ebola virus from pigs to non-human primates. Sci Rep. 2012;2:811. doi: 10.1038/srep00811. Epub 2012 Nov 15.