Mystery Rays from Outer Space

“Batrachia”, by Ernst Haeckel (Kunstformen der Natur, 1904)

There’s a constant viral assault on us humans, as there is on just about all other species. We as a species have to contend not only with the vast pool of human pathogens, those viruses that constantly circulate among humanity; but also with the continual probes on our defenses from other viruses, viruses that normally infect other species.  All of us are exposed to these on a regular basis: Dog and cat viruses, mouse viruses, crow and pigeon viruses, bat viruses, not to mention the ocean of insect and fungus and amoeba and plant viruses.

Almost all of these assaults don’t even scratch our defenses.  The viruses can’t even enter our bodies, and if they do then they can’t enter our cells, and if they do they can’t replicate in our cells, and if they do then they can’t  …

Most viruses, in other words, can’t effectively jump species.  Even when they do, they’re usually not well adapted to the new species, and they can’t establish a productive chain of infections. Even if they cause a disease, they burn themselves out, infecting fewer and fewer individuals each round of infection, until they disappear.

But every so often, in a tiny minority of cases, the virus does get a foothold.  This is one of the ways that “emerging infections” get started.  It covers things like HIV, SARS, parvovirus of dogs, Ebola, and of course the new H1N1 swine-origin influenza virus (SOIV), among many others.

Why did these guys take off, when so many other viruses failed? Why did SOIV infect people last year, while decades of exposure to pigs and swine H1N1 influenza viruses didn’t lead to earlier pandemics?  Basically, we don’t know, and we’d really, really like to know, so we have a chance of predicting the next SOIV or HIV before it’s a pandemic.

OK, so that explains why I’ve written a fair number of posts here on species-jumping in viruses (here, here, here, here, and here), and partly explains why I want to mention a new paper from Bertram Jacobs‘ lab¹.  (The rest of the reason is, as always, that I just think it’s  cool.)  I’m not sure why Jacobs has done this particular project, because he’s more of an interferon guy, but he’s looked at the origins of ranaviruses and finds evidence for lots of species shifts in their history.

“The Smooth Terrapin (Emys terrapin)”, by James Dekay (Zoology of New York; or, The New York fauna, 1843)

Ranaviruses are probably best known as frog viruses, but they infect a bunch of cold-blooded animals — fish, frogs, salamanders, turtles, and so on — and several of them are causes of emerging infectious disease (as I discussed last time I talked about ranaviruses, here).  Jacobs’ group looked at about a dozen of them whose genomes are completely sequenced², and tried to put together their evolutionary history, which turns out to involve all kinds of cross-species jumps:

…we hypothesize that the most recent common ancestor of the ALRVs was an ancestral fish virus …  Both of these hypotheses suggest that for the majority of evolutionary time vertebrate iridoviruses were confined to fish, and much more recently, there appear to have been at least three species jumps, from fish to frogs, from fish to salamanders, and from frogs to reptiles, and perhaps as many as four species jumps, including a jump from tetrapod amphibians back to fish. It is tempting to speculate that activities associated with human harvesting of aquatic organisms during the past 40,000 years led to the more common recent jumping of ranaviruses among aquatic organisms.¹

(My emphasis) They don’t offer any specific reasons why the ranaviruses should be able to leap from species to species like the chamois of the Alps, but they do make the general point that these viruses tend to be rather promiscuous to start with.  Not only are closely-related viruses able to infect different hosts, but even the same viruses often are able to infect a wide range of species; the fish virus they sequenced in this paper, epizootic hematopoietic necrosis virus, can infect a half-dozen different species of fish.  They raise an interesting comparison:

In addition, the ability of this group of viruses to infect such a wide variety of host species suggests that more host shifts are likely. Therefore, it is important that we understand more of the evolutionary traits of this unique group of viruses, as there is no other closely related group of viruses that infect such a broad group of hosts, with the possible exception of the orthomyxoviruses.¹

Orthomyxoviruses, of course, include influenza viruses, which notoriously infect humans, pigs, ducks, chickens, wild waterfowl, horses, and dogs; and you’ll recall all the reports during the epidemic phase of SOIV of the virus infecting all kinds of other pets and domestic animals.  Influenza viruses are apparently evolving at an even faster pace than the ranaviruses, and experimenting with even more species; but there may be lessons for us (as influenza hosts) in the ranaviruses.

1. Jancovich, J., Bremont, M., Touchman, J., & Jacobs, B. (2009). Evidence for Multiple Recent Host Species Shifts among the Ranaviruses (Family Iridoviridae) Journal of Virology, 84 (6), 2636-2647 DOI: 10.1128/JVI.01991-09[↩][↩][↩]
2. Including epizootic hematopoietic necrosis virus, whose genome they sequenced themselves[↩]

Most of us don’t think much about yellow fever nowadays. There are still a couple hundred thousand cases, and some 30,000 deaths, each year, but almost none are in the first world. Out of sight, out of mind.

But this indifference is new. Until the beginning of the 20th century, yellow fever ran rampant, and was one of the most dreaded of all diseases. Epidemics of yellow fever in New York, Philadelphia, Memphis, and New Orleans killed tens of thousands. There’s a WHO manuscript [pdf link] on yellow fever that lists these and many more outbreaks – page after page of fine-print dates and deaths.

Those who have not lived between Cancer and Capricorn can well fail to conceive readily the sensation of numb, chill dreariness which steals on all hearts, when the news spreads, from mouth to mouth, that Yellow Jack has once more come. … at last it is admitted on the housetops, as well as whispered in the closet, that the deadliest, most awe-inspiring of the plagues of the equatorial regions has obtained admittance within our borders.

–”Yellow Jack“, in Cornhill Magazine, 1892¹

As I noted earlier, Carlos Finlay made the original suggestion that yellow fever was a mosquito-borne disease in 1881;² in English, in 1886. ³ Walter Reed and his team confirmed this in 1900, and the discovery was seized on at once.

“Shows the Distribution of the Principal Mosquitoes of New Orleans”. Dark squares represent Stegomyia fasciata, the major carrier of yellow fever 4	Yellow fever cases in New Orleans, 1905. “The infected blocks are most numerous in the old, Italian, quarter of the city.”

(This post was mainly an excuse to post those maps.⁵  Click for larger versions.)

New outbreaks were checked with enthusiastic mosquito control.

In a few days with very little opposition, sixty to seventy thousand cisterns had been screened in order to prevent the breeding of the Stegomyia fasciata. Mosquito nets became more than ever the rule …

–Yellow Fever Prophylaxis in New Orleans, 1905⁵

The virus itself was was isolated in 1927 and the vaccine, made in 1937 by Max Theiler, turned out to be extremely effective; but even before that,  the understanding that mosquitoes were the carriers allowed great strides in reducing the disease.  Not just in the USA, but throughout the Americas:

Havana and Cuba freed from fever by Gorgas, who organized anti-mosquito measures, 1901-1902; example followed in Rio de Janeiro and Vera Cruz, 1903-1909; Panama Canal Zone successfully protected by same methods, 1904-1906 …  intensive campaign, 1918-1919, under Connor eliminated disease from Guayaquil, the chief endemic centre …

–”Yellow Fever in Retreat“, 1922⁶

1. Cornhill Magazine
   New Series, Vol. XIX, July to December 1892
   Smith, Elder, & Co.
   15 Waterloo Place, London [↩]
2. C. Finlay (1881). El mosquito hipoteticamente considerado como agente de trasmislon de la flebre amarllla An. de la Real Academia de ciencias med. de la Habana, 18, 147-169[↩]
3. C. Finlay (1886). Yellow Fever, its transmission by means of the Culex mosquito Am. Journ. Med. Sci., 92, 395-409[↩]
4. If I follow this right — I’m neither an entymologist nor an entomologist — Stegomyia fasciata was subsequently renamed Stegomyia aegypti, then Aedes agyptyi, and now (since 2005) is properly is officially called Stegomyia aegypti once again but usually, if not always, with “Aedes” in brackets to clarify. There was also, maybe, a point at which it was Stegomyia calopus, unless that was something else.[↩]
5. Yellow Fever Prophylaxis in New Orleans, 1905
   Rubert Boyce
   April, 1906
   Published for the Committee of the Liverpool School of Tropical Medicine
   by Williams & Norgate
   14 Henrietta Street, Covent Garden, London[↩][↩]
6. Current History
   A Monthly Magazine of the New York Times
   Volume XVI, April-September, 1922[↩]

Man chasing rabbit (From “Fliegende Blätter”, Munich, 1889)

Everyone knows about rabbits in Australia. Introduced in the mid-1800s, they multiplied ridiculously and ate their way across the country, leaving barren devastation behind them.

Myxomavirus, a poxvirus that originated in South America, was introduced in the early 1950s and temporarily controlled the rabbit population, cutting their numbers by 85% (to a mere hundred million rabbits); but the rabbits evolved some resistance, and the virus evolved somewhat reduced virulence, and after about 15 years the rabbit population started to build up again. (I’ve talked about myxomatosis and rabbit control here and here.)

Myxomavirus isn’t a natural pathogen of European rabbits; its natural hosts are American rabbits, in which it causes a much more mild disease. It’s a virus that jumped into a new species, was very virulent in that new species, and then became less so over 15 years or so of transmission in the new species.

Myxomavirus is often used as an example of a virus that evolves toward avirulence, with the message usually being that this is the usual path of evolution. For example, you’ll see comments like, “Typically, viruses that rapidly kill their host have a very short history, as they rapidly run out of places to reproduce.” As I’ve tried to point out several times (see the myxomavirus links above), this isn’t true; pathogens in general evolve toward improved transmission, not reduced virulence. In many cases, reducing virulence does enhance transmission, but it’s not the only path. And myxomavirus doesn’t even support the claim all that well, given that even the “low-virulence” strain that’s out there now still has a mortality rate about the same as Ebolavirus, or smallpox.

OK, hold that thought.

Extermination of rabbits in California (From “The Picture Magazine,” 1894)

Myxomavirus worked well to control rabbits for a while, then became less effective. In 1995, a new virus was introduced into Australia and New Zealand,¹ a calicivirus that causes Rabbit Hemorrhagic Disease, called (with stunning originality) Rabbit Hemorrhagic Disease Virus (RHDV). Where did RHDV come from?

Basically RHDV is the opposite of myxomavirus. The parent of RHDV is a natural virus of European rabbits, but it causes little or no disease. RHDV is a natural mutation of this virus, and it has very high virulence – the opposite of the viruses-evolve-to-low-virulence claim. Even with the help of Australian farmers, RHDV is highly successful. It spread around the world in the mid-1980s after first appearing in Chinese rabbits in 1984.

One of the most intriguing aspects of RHDV evolution is that this virus appears to have maintained its very high virulence during the 25 years since it emerged. At face value this suggests that virulence is adaptive for transmission. ²

Rabbit’s eye (Max Brödel, 1932)

In fact, it’s been believed that the virulent RHDV is such a successful mutation that it arose several times, independently, from the mild parent virus. (Compare to the feline infectious peritonitis story, where the prevalent explanation for the appearance of a virulent FIP infection is that innocuous gastrointestinal coronaviruses, that are widespread in cats, mutates to form the virulent form; and this mutation is independent for each cat, rather than arising once and then spreading.)

At least, that’s been the established explanation, but a recent paper² shows that a couple aspects of that aren’t correct. The authors looked at genome sequences of many rabbit caliciviruses (the mild parent calicivirus as well as the virulent RHDV) to track its origins and spread. RHDV is indeed a recent mutation from a mild parent virus; that much is correct. But RHDV probably only arose once, not multiple times, and the origin of RHDV was well before 1984) when it was identified as a disease.

The independent-mutation hypothesis was based mainly on finding RHDV-related viruses circulating in Europe in the 1950s:

Prior phylogenetic work led to suggestions that RHDV with sequences closely related to those that emerged from China in 1984 were circulating harmlessly in the United Kingdom and other European localities during the 1950s; hence, it was suggested that virulence emerged at least twice during the late 20th century: once in Europe and once in China. ²

But Kerr et al looked more closely at these early isolates, and don’t think they’re real:

… we show here that the sequences from the 1950s and 1970s from the United Kingdom appear to be modern contaminants: given the rate of RHDV evolution documented here and that of RNA viruses more generally, these early RHDV sequences are expected to be far more divergent from their modern counterparts. ²

(Compare to the influenza database, where there also seems to be a significant level of mis-identified virus.)

So RHDV probably only originated once, which is a little more reassuring than the notion that this high virulence is so easy to achieve that it can appear many times over a short period. Did the original mutation appear around 1984, when the disease was noted? The authors identified 4 distinct strains of RHDV and noted:

A common feature of all of these groups is that many lineages likely originated during the 1970s, suggesting that there was a period of viral radiation at this time…. Crucially, this also means that there were already multiple separate lineages of RHDV before the documented emergence of RHD in China in 1984.  … This implies either that high virulence evolved multiple times in multiple viral lineages close to 1984 or (more plausibly) that virulence emerged earlier in the 20th century but the disease was not documented until 1984 when the trade in rabbits provided the opportunity for RHDV to spread from an established, but apparently cryptic, transmission cycle. … Therefore, we propose that the most likely scenario is that virulent RHDV strains evolved once, early in the 20th century, but were not detected until 1984. ²

(My emphasis) This seems, at first glance, surprising. RHDV kills almost all of the rabbits it infects. Wouldn’t you notice it if all your rabbits suddenly fell over dead? How could RHDV circulate for years or decades without being detected? Kerr et al make some points about the nature of the disease (it can infect very young kits without killing them, for example), but also comment:

Given the difficult sociopolitical conditions in China and neighboring countries in the first half of the 20th century, it is plausible that a virulent disease in rabbits was able to evolve in this region without leaving a clear record. ²

The pandemic swine-origin H1N1 probably has been circulating in swine for quite a while (years? decades?) without being picked up, and probably circulated in humans in Mexico for months before it was detected there. It’s pretty easy to believe that rabbits in China during the Cultural Revolution didn’t get as much attention as pigs in the US in the 2000s.

So, an interesting story in its own right, especially thinking about evolution of virulence in pathogens; and also, a story that probably reflects some important lessons for human health.

1. It wasn’t supposed to be introduced then; it was penciled in for few years later, after more study, but somehow it jumped from the island where it was being studied to the mainland. The usual explanation is “via insects”, but of course one has to wonder if some Australian farmers didn’t help the insects along some. As I recall from the news reports at the time, the “accidentally introduced” virus spread throughout Australia very fast, almost as if dead rabbits were being carted around by car or something.[↩]
2. Kerr, P., Kitchen, A., & Holmes, E. (2009). Origin and Phylodynamics of Rabbit Hemorrhagic Disease Virus Journal of Virology, 83 (23), 12129-12138 DOI: 10.1128/JVI.01523-09 [↩][↩][↩][↩][↩][↩]

Despite the fact that the recent pandemic was the best studied and recorded to date, the knowledge gained will probably have little predictive value for the next pandemic, either in qualitative or quantitative terms.

–Communicable Diseases and Epidemics
Martin M. Kaplan
Bulletin of the Atomic Scientists, June 1960

Kaplan  was referring to the 1957-58 influenza pandemic, but the sentence could have been taken from many places in 2009. In any case, Kaplan was pretty much correct; the next three pandemics after that were quite different from the 1957-58 pattern, and quite different from each other.

Edit: I’ve updated the table to reflect the CDC’s numbers for age distribution of infection, which I didn’t see first time around.  Thanks to Marcello Pucciarelli for the link.  The original version of this post, containing my guesswork on the distribution, is still available here. Using the more accurate numbers has very little effect on my overall conclusions.

I’ve been expecting a resurgence of swine-origin influenza virus (SOIV) in North America for a while now, and it hasn’t happened. The virus is still out there, still infecting a few thousand people per week, but there hasn’t been a third large-scale wave of virus transmission. That’s different from the 1918 and 1957 pandemics (see here for details). What’s different this year?

Of course there are tons of things that are different this year, but I recently got around to doing something I should have done a while ago: Trying to estimate what proportion of Americans are now immune to SOIV. I’ve seen estimates that around 40% of the population should now be immune. I get roughly the same number (slightly higher, closer to 50%, because those estimates didn’t take into account pre-existing immunity to SOIV), but I think there’s an important point that might be missed in this: Most of the immunity might be clustered in the two most susceptible populations (children and the elderly), with two-thirds to three-quarters of them being immune.

There are three ways someone could be immune to SOIV. They could have been exposed to a related virus, some time in the past, and have developed a long-term immunity. They could have been infected with SOIV, somewhere in the first or second wave. Or, of course, they could have been vaccinated.

Numbers for each of those are available. They’re more or less approximate; not all the sources break down age groups in exactly the same way, for one thing, and I don’t have precise numbers for everything.¹ I’ll try to flag places where I’m especially guessing, and it’s entirely possible that this I’ve made some obvious, basic mistakes in here, since this has been written in the interstices of cleaning our house for Chinese New Year (a Herculean task) and hosing down the kids to get them ready for the party at YongHui’s this evening.  If so, let me know and I’ll try to correct them.

We need to break immunity down by age, because pre-existing immunity to SOIV was strongly age-dependent. (That’s presumably why the virus was strongly biased to infecting younger people this year.) For the demographics of the US I’m using the 2008 census data. All this is summarized in the table below, and my explanations follow.

1. Pre-existing immunity. A paper in New England Journal of Medicine last year² found that 4% of children, 6% of young adults, and 34% of older adults (born before 1950; I’m going with 65 years old as the dividing line just to make it easier to compare to other data) were already immune to H1N1. That’s more or less consistent with other studies I’ve seen, so let’s go with that.

2. Infection. The CDC estimates that somewhere over 55 million people in the US were infected in the first and second wave of SOIV, and gives approximate age distribution here. I’ve used the mid-point of their range estimates, so it’s possible that significantly more people were infected.  This works out to a quarter of US children being infected with SOIV,  which is broadly consistent with measures elsewhere — for example, a recent Lancet paper³ that estimated that about a third of children in the UK were infected.

3. Vaccination. The CDC’s Anne Schuchat’s Feb 5 press conference was very useful for this. The CDC has estimated that somewhere over 70 million people in the US have been vaccinated. Schuchat gave two further figures: Some 37% of children, and about 21% of adults have been vaccinated. These figures come from two different sources — a CDC survey and a Harvard survey respectively — so they may not be directly comparable, and I don’t know the breakdown in adults (young adults vs. elderly) but these figures do work out to about the right total, around 80 million people. Probably a little high, but not by much.

Another source of fuzziness is how much overlap there is between infected people and vaccinated people. It’s probably safe to say that most vaccinated people were not subsequently infected, but it’s quite possible that people were infected (perhaps with no symptoms, which we know happened quite frequently) and were subsequently vaccinated. ⁴ I’ve tried to adjust for this by assuming that half of infected people didn’t know it, and went on to get vaccinated, ⁵ as well as subtracting the proportion of people who were already immune (who presumably had no way of knowing that).  That’s the “Vaccinated uninfected” column.  Or, I’ve assumed  no overlap (just plain “Vaccinated”), to get an approximate range of immunity out there.

Including “Vaccinated uninfected” gives the “Number immune (low)”; assuming that all vaccinated were not infected gives the “Number immune (high)”.

And my conclusions are that:

• Over half the US population as a whole is now immune to the new SOIV.
• As many as three-quarters of the elderly and two-thirds of the children — the most important populations as far as flu is concerned — may be immune.
• Between a third and about half of this immunity was due to vaccination.

That level of immunity is probably enough to impact virus transmission drastically. In the early waves, if a child was infected then virtually every child she contacted in school or on the playground would be susceptible. Now only one in three of them are potentially infectable. I’ll have to spend some time looking at the models of influenza spread but I think that considering that the SOIV was not spectacularly infectious anyway, this level of population immunity is easily enough to prevent the third winter wave of disease I was expecting to see.

(I am particularly curious about modeling the impact of vaccination. Without vaccination would there have been a third wave? My guess is that there would have been, but that’s just a guess.  Update for clarification: Vaccination rates were highest in children. Without vaccination only about 25% of children would be immune — vaccination therefore doubled or tripled the amount of immunity in this critical population, and I think SOIV would have resurged in schools in winter without this intervention.)

What’s more, this level of immunity, especially in the apparent absence of the usual seasonal flu strains, has important implications about influenza over the next few years, but this post is already too long, so maybe I’ll talk about that some other time.

1. Probably quite accurate numbers are available, but not to me. Or, at least, not without a lot more work.[↩]
2. Hancock, K., Veguilla, V., Lu, X., Zhong, W., Butler, E., Sun, H., Liu, F., Dong, L., DeVos, J., Gargiullo, P., Brammer, T., Cox, N., Tumpey, T., & Katz, J. (2009). Cross-Reactive Antibody Responses to the 2009 Pandemic H1N1 Influenza Virus New England Journal of Medicine, 361 (20), 1945-1952 DOI: 10.1056/NEJMoa0906453[↩]
3. Miller, E., Hoschler, K., Hardelid, P., Stanford, E., Andrews, N., & Zambon, M. (2010). Incidence of 2009 pandemic influenza A H1N1 infection in England: a cross-sectional serological study The Lancet DOI: 10.1016/S0140-6736(09)62126-7[↩]
4. This is another significant potential source of error, I think.[↩]
5. That is, I’ve subtracted that proportion of people from the vaccinated totals.[↩]

Vesicular stomatitis virus (VSV), like the related rabies virus, is a bullet-shaped virus.  Hong Zhou has just added VSV to his collection of cryo-electron microscopy virion structures,¹ and as always with viruses, it’s just gorgeous.

“Architecture of the VSV virion. … A montage model of the tip and the cryo-EM map of the trunk”1	“A plausible process by which the nucleocapsid ribbon generates the virion head, starting with its bullet tip.”1

1. Peng Ge, Jun Tsao, Stan Schein, Todd J. Green, Ming Luo, & Hong Zhou (2010). Cryo-EM Model of the Bullet-Shaped Vesicular Stomatitis Virus. Science, 327 (5966), 689-693 DOI:10.1126/science.1181766[↩][↩][↩]

Every so often — not often enough — I run across a paper that’s so ridiculously ingenious that it just makes me laugh with pleasure.

Ladies and gentlemen, a round of applause, please, to Shou-Wei Ding, of the Center for Plant Cell Biology at UC Riverside, for his Rube Goldberg-esque brilliant technique for identifying new viruses. ¹

Background: Small interfering RNA (siRNA, ² or RNAi) is pretty well known nowadays, especially since three of its discoverers were given Nobel Prizes a few years ago. These small RNAs are found in most eukaryotes — plants, insects, worms, as well as birds and mammals. In fact, siRNA was first discovered in plants and then was widely used in insects and worm research well before it was shown in mammals. In mammals, we tend to think of small RNAs as having regulatory functions. In plants, insects, and worms, there are certainly regulatory siRNAs, but siRNAs are also used as an important part of their antiviral immune response. (Unsurprisingly, plant and insect viruses themselves have defenses against these siRNAs, by producing anti-siRNA genes.)

siRNAs are small (duh), maybe 20-30 bases long. Without going into mechanisms more than necessary, they recognize specific sequences of their target RNA (by being complementary to the target). When they bind to their target, they cause that target RNA to be chopped up into small pieces. Some of those small pieces can then act as new siRNA and the cycle continues.

Ding’s group reasoned that (1) if insect viruses are being attacked by siRNA, and (2) the viruses are then chopped up into new siRNA, then (3) all you have to do is piece together the siRNA, to recover the virus sequence.

Seems sort of obvious now that I put it together that way, but I would have said that there’s no way it would work — not enough coverage, I would have said, you’d only see a tiny fraction of the genome. Even if there was enough coverage, how would you find the virus pieces in the huge pool of other siRNAs? And even if you could do that, how would you piece them together? It would be like a jigsaw puzzle where every piece was just a tiny snippet of blue sky.

But, wonderfully, it actually worked for Ding’s group. Not only did they identify viruses that they knew should be there, they also pulled out five brand-new viruses out of their insect cells:

In this study, we found that viral small silencing RNAs produced by invertebrate animals are overlapping in sequence and can assemble into long contiguous fragments of the invading viral genome from small RNA libraries sequenced by next-generation platforms. Based on this finding, we developed an approach of virus discovery in invertebrates by deep sequencing and assembly of total small RNAs (vdSAR) isolated from a host organism of interest. Use of this approach revealed mix infection of Drosophila cell lines and adult mosquitoes by multiple RNA viruses, five of which were previously undescribed.

“Virus discovery in OSS cells by viral genome assembly from sequenced viral piRNAs of 25–30 nucleotides in length after viral siRNAs were removed.”

The ability to piece these tiny fragments together is a spin-off of the new genome sequencing platforms, which by their nature make very short reads that have to be computationally stitched back together. Most of these new platforms make slightly longer fragments than siRNA size, but they’re in the same ballpark and I guess the same approaches work.

As far as the coverage, they didn’t find 100% of the genomes, but they got a really surprisingly high fraction back — 80% to 95% or more of the various viruses.

Not only that, they got really new viruses:

As a result, none of the four viruses could be assigned into an existing virus genus. This suggests that vdSAR is capable of discovering viruses that are only distantly related to known viruses.

Giving credit where it’s due, another group recently, and independently, used a similar approach to identify viruses in sweet potatot plants³ but I didn’t notice that article until Ding pointed it out.

Whether or not this technique proves useful in the long run, it’s just so ingenious that I want it to succeed.

1. Wu, Q., Luo, Y., Lu, R., Lau, N., Lai, E., Li, W., & Ding, S. (2010). Virus discovery by deep sequencing and assembly of virus-derived small silencing RNAs Proceedings of the National Academy of Sciences, 107 (4), 1606-1611 DOI: 10.1073/pnas.0911353107[↩]
2. I’m going to call all of the various types of small RNAs “siRNA” here, but that’s just shorthand, there are different subclasses that I won’t go into[↩]
3. Kreuze, J., Perez, A., Untiveros, M., Quispe, D., Fuentes, S., Barker, I., & Simon, R. (2009). Complete viral genome sequence and discovery of novel viruses by deep sequencing of small RNAs: A generic method for diagnosis, discovery and sequencing of viruses Virology, 388 (1), 1-7 DOI: 10.1016/j.virol.2009.03.024[↩]

I didn’t post anything about the recent study¹ showing that handwashing + face masks reduces influenza spread, because other blogs covered it fairly extensively (for example, here’s Avian Flu Diary’s commentary). Here’s another study giving a common-sense check:

… in a household setting, simple, readily available products such as 1% bleach, 10% vinegar and 0.01% washing up liquid all make convenient, easy to handle killing agents for influenza virus A/H1N1. These findings can be readily translated into simple public health advice, even in low resource settings. The public do not need to source more sophisticated cleaning products than these.

–Greatorex, J., Page, R., Curran, M., Digard, P., Enstone, J., Wreghitt, T., Powell, P., Sexton, D., Vivancos, R., & Nguyen-Van-Tam, J. (2010). Effectiveness of Common Household Cleaning Agents in Reducing the Viability of Human Influenza A/H1N1 PLoS ONE, 5 (2) DOI: 10.1371/journal.pone.0008987

(My emphasis) Their figures show that these common solutions almost immediately reduced the numbers of virus from between 1 and 100 million at the start, to undetectable levels (less than 200). Hot water, not surprisingly, didn’t work.

They also added that “branded anti-bacterial wipes and anti-viral tissues were encouragingly effective at inactivating the virus“, so if you’d rather buy something expensive, go ahead.

1. Aiello, A., Murray, G., Perez, V., Coulborn, R., Davis, B., Uddin, M., Shay, D., Waterman, S., & Monto, A. (2010). Mask Use, Hand Hygiene, and Seasonal Influenza?Like Illness among Young Adults: A Randomized Intervention Trial The Journal of Infectious Diseases, 201 (4), 491-498 DOI: 10.1086/650396[↩]

OK, last time I thought H1N1 influenza was coming back (just after Christmas) it turned out to be just a blip.  But I notice that according to Google Flu Trends, 30 states are showing increases in flu activity this week, compared to last week. Mostly very small increases,¹ but 10 of the states have shown a sustained increase (at least two weeks of increasing numbers). Over the past two weeks, Alabama, Kansas, Louisiana, Oregon, and Utah have each shown a 10-20% increase in flu activity.

This is pretty much the time that flu season normally begins, but there’s been very little evidence of the normal seasonal flu strains circulating this year, so odds are these cases are almost all H1N1 swine-origin influenza virus.

Just saying.

1. None are more than about 10% higher than last week[↩]

Last week, the Effect Measure blog¹ talked about a paper that offered a new way of treating influenza.² Briefly, the approach is to attack the virus by treating the host cell: Eliminating host functions that the virus requires, but that the host cell does not.

The authors of the paper commented that “targeting host cell determinants temporarily dispensable for the host but crucial for virus replication could prevent viral escape,” and Effect Measure observed that “It’s not obvious to me why the virus can’t as easily mutate in ways to adapt to a missing “office tool” as to a drug that affects an important viral function.” In the comments, I said:

I think the fundamental difference is that in the latter case, the virus needs just modify an already-present function; but if the tool is missing altogether, the virus would have to develop a whole new function from scratch, an altogether more difficult task.

That’s not to say that a virus could not do it — we see examples of this all the time, with viruses that have co-opted host functions and in some cases even host genes. But the process is usually much slower; we tend to recognize those events in hindsight, whereas we can often watch viral genes adapting to drugs in real time.

I still think my explanation is generally true, but here’s a counterexample, of what appears to be a virus evolving a brand-new function in just 5 weeks.

Background: Mammals have what seems to be a general defense against retroviruses like HIV. Several members of the APOBEC family of proteins are anti-retroviral;³ they force widespread mutations into the HIV genome, so many mutations that the virus can’t replicate or produce normal proteins.⁴ The reason HIV is able to replicate in spite of the APOBECs, is that HIV has in turn an anti-APOBEC protein, vif, that causes rapid destruction of several APOBECs. (I’ve mentioned this before, here, here, and here.)

Predicted vif structure (Zhang et al.)

Variants of HIV that don’t have vif (either natural or artificial) can replicate pretty much normally in cells that don’t produce APOBECs; but they’re dead in cells that do have APOBECs, and their natural targets for infection do have APOBECs. So HIV is pretty much absolutely dependent on vif for its life-cycle.

There’s a lot of interest in trying to use this fact as an anti-HIV treatment. If there was a safe, effective anti-vif drug, then the APOBECs that are normally present could go ahead and destroy the virus. Now, we know from experience with other anti-HIV drugs that the virus could probably mutate vif to avoid this hypothetical drug, but let’s say it couldn’t. Let’s say the drug was completely effective in blocking vif, and there was no way for HIV to build a drug-resistant vif. Would the virus be completely helpless, or would it be able to develop a whole new anti-APOBEC function from scratch?

At least in a specific and somewhat limited set of conditions, that’s just what happened. Hache et al⁵ took a vif-deleted virus and tossed it into cells that have APOBEC3G. For several weeks, as you’d expect, there was almost no virus recovered (because without vif, the virus was destroyed by the APOBEC3G). But (see the figure to the left here; click for a larger version) after about 45 days, in 3 of the 48 cultures, virus abruptly started to grow again.

“Highlights of the long-term spreading-infection experiments for Vif-deficient viruses on vector-control- or APOBEC3G-expressing CEM-SS cell lines. Of the cultures, 45/48 showed no virus replication on APOBEC3G-expressing cells (flat lines not graphed).” 5

These new viruses still didn’t have any vif, but they were pretty much resistant to APOBEC3G — they had developed a brand-new function that conferred resistance to APOBEC3G. This new function behaves quite differently from vif. For one thing, vif protects against several different members of the APOBEC family, while the new variants were only resistant to APOBEC3G (they were still susceptible to APOBEC3F). And it took two simultaneous changes in the viruses for this to work: “Virus replication was only detectable after two mutations appeared: a noncoding A200T(C) transversion and a Vpr null mutation.“

The mechanisms underlying this aren’t quite clear, and it’s really mysterious why getting rid of vpr would help make HIV resistant to APOBEC3G (vpr is a fairly mysterious protein in its own right, so it doesn’t offer a lot of handles to work it out). Anyway, although the authors did offer a number of possible explanations, that’s not really what I wanted to talk about. The point I wanted to make is that viruses can acquire new functions out of nothing, as well as modifying already-present functions.

Co-expression of Vif and APOBEC3G in HeLa cells

Having said that, I think this actually does support my answer to some extent. Would this sort of resistance actually arise in vivo? Remember the question the authors were looking at originally: If we have an anti-vif drug, will resistance to it quickly arise? And if we look at the characteristics of the resistance in this artificial system, it’s actually somewhat encouraging:

• The resistance took quite a while to pop up — several weeks, anyway ⁶
• The resistance was rare even over that timescale. Only three of the 48 cultures threw out resistance variants. Each culture was infected with thousands of viruses originally,⁷ so you could say that the rate was lower than 3 in 400,000⁸

(Both of these are sort of what you’d expect from variants that require multiple mutations for resistance. Single mutations occur very frequently, but multiple mutations are exponentially⁹ less frequent.)

• The resistance was partial. These viruses were only resistant to APOBEC3G. In a natural infection, these resistant viruses would probably still be killed, because they’re still susceptible to APOBEC3F

So: Yes, viruses can develop new functions, but it’s probably still fair to say they’re not as adept at this as at modifying existing functions.

1. Which you should be reading, if you’re not already[↩]
2. Karlas, A., Machuy, N., Shin, Y., Pleissner, K., Artarini, A., Heuer, D., Becker, D., Khalil, H., Ogilvie, L., Hess, S., Mäurer, A., Müller, E., Wolff, T., Rudel, T., & Meyer, T. (2010). Genome-wide RNAi screen identifies human host factors crucial for influenza virus replication Nature DOI: 10.1038/nature08760[↩]
3. Sheehy, A. M., N. C. Gaddis, J. D. Choi, and M. H. Malim. 2002. Isolation of a human gene that inhibits HIV-1 infection and is suppressed by the viral Vif protein. Nature 418:646-650.[↩]
4. Mangeat, B., P. Turelli, G. Caron, M. Friedli, L. Perrin, and D. Trono. 2003. Broad antiretroviral defence by human APOBEC3G through lethal editing of nascent reverse transcripts. Nature 424:99-103[↩]
5. HACHE, G., SHINDO, K., ALBIN, J., & HARRIS, R. (2008). Evolution of HIV-1 Isolates that Use a Novel Vif-Independent Mechanism to Resist Restriction by Human APOBEC3G Current Biology, 18 (11), 819-824 DOI: 10.1016/j.cub.2008.04.073[↩][↩]
6. How does this compare to the time it would take HIV to pop up resistant variants to one of the drugs in HAART therapy?  Anyone know? Bueller? Anyone?[↩]
7. It’s not clear from their methods exactly how many, but 10,000 seems like a reasonable guess[↩]
8. That’s not really a valid interpretation but it offers an upper limit, anyway[↩]
9. Exponentially? Geometrically? Anyway, “much less”[↩]