Tag Archives: transmission

H5N1 Ferret Transmission Experiment Published

At last, one of the papers investigating H5N1 influenza transmission in ferrets has been published in the journal Nature yesterday.  To recap the controversy briefly:  news of experimental studies investigating transmissibility of avian H5N1 influenza hit the news this fall, igniting a fierce debate about biosecurity, “dual use” research, and the damage that censorship can have on scientific advancement.  An advisory group, the NSABB, recommended partial censorship of the data, perhaps believing that redacting specific data from the publications would prevent information on how to generate a highly pathogenic mammalian transmissible virus from getting into the hands of bioterrorists or others incapable of handling such viruses safely.  However, after some new data or clarification of data presented in a revised version of the submitted manuscript, the NSABB recommended publication in full.

Avian H5N1 influenza virus has caused sporadic infections in humans who have close contact with infected animals.  Human-to-human transmission has not been observed.  But could an H5 virus mutate or reassort, allowing human-to-human transmission?

One thing that has been lost in this whole controversy is that this study is actually a great demonstration and application of evolution.  How does a virus switch to a new host and transmit efficiently between individuals?  Start with a diverse population that varies in a particular trait (in this case, ability to bind the human receptor).  Put it through selective pressure. This occurred in several steps: first, select for viruses that can bind the human receptor in vitro.  From those that bind, select ones that can efficiently replicate in the respiratory tract of the animal.  Finally, take the efficient replicators and allow for transmission.  At each step of selection, mutations that naturally occur during viral replication further diversify the population resulting in variants that possess the desired property.  Those variants get selected for the next experiment.

This study focused on one particular influenza virus protein, hemagglutinin (HA).  HA is on the surface of the virion and is what the virus uses to attach to the host cell, the first step in viral infection.  HA of human influenza viruses bind a sugar on the surface of human cells, which is slightly different from that found in the avian respiratory tract.  Avian viruses, of course, bind to the form found in birds.  H5 shows a strong preference for binding the avian receptor, so Karaoka et al were interested in finding out if H5 could change to recognize the human receptor, allowing more efficient transmission between mammals.

To address this, they began with a mutagenesis technique to introduce random mutations in the globular head (the receptor binding part) of HA, then used an in vitro approach to select for mutants that bound the human receptor (selection step 1).  Through this process they identified three H5 variants that gained the ability to bind the human receptor while maintaining the ability to bind the avian receptor, and one that switched specificity completely to the human receptor.  Several of the mutations identified in this study had already been shown in previous studies to be important in receptor specificity.

To test if the variant H5s conferred binding to human receptors in vivo, sections of human tracheal tissue were exposed to the viruses and only two were able to infect (selection step 2).  This suggests the virus can infect human epithelium of the upper respiratory tract.

Next, the two remaining variants were used to infect ferrets.  Both replicated in ferret respiratory tracts, but one replicated to higher levels.  When they sequenced the virus that they isolated from that ferret, it was different from what they had put in: a new mutation had appeared.  This new mutation presumably confers the property of better replication in the ferret respiratory tract, so it outgrew the original input virus (selection step 3).

Using this new virus (now with a total of 3 mutations in H5), transmission was tested in ferrets.  Compared to the original H5, which did not transmit via aerosol, the 3-mutation variant did transmit, although between only 2 of 6 animal pairs.  Again, they sequenced the virus that was present in the contact animals and found that it was different than what had gone in to the inoculated animals.  Yet another mutation had appeared (selection step 4).  This additional mutation appears to enhance transmission:  the new virus, now with 4 mutations, transmitted more quickly and between more pairs of animals than the 3-mutation virus.  Although the virus can transmit, none of the infected animals died, but they did show pathology at the site of infection.

So what does this all mean? The best model available for influenza transmission studies is ferrets.  Ferrets aren’t humans, so its important to keep in mind that this is a model that helps us understand what viral or host factors are involved in aerosol transmission in these mammals, and maybe, but not necessarily, in humans.  Since we don’t know what is necessary for human-to-human transmission, it is valuable to have an animal model to give us some ideas of what to look for.  It can provide some good hypotheses on what mediates transmission in humans, which would then have to be further tested.  Obviously, specificity for the human receptor is necessary, but the mutations identified tell us more than that.  Mutations that change specificity are not sufficient for transmission.  It turns out that those mutations also decrease the stability of the HA protein.  The additional mutations acquired through the selection steps compensate for that, and enhance stability.   So now we know that HA stability is important in influenza transmission.  Between ferrets.  That’s probably true for humans too, but it would need to be tested.

If you are still reading, you are obviously procrastinating, and are probably avoiding studying for your final exam.  But here are some more thoughts on the controversy overall.  This is a really interesting paper, with nothing particularly frightening or worrisome about it.  Certainly not any more so than other papers doing similar work that were published without so much controversy.  If “dual use” research needs to be regulated, it needs to be done before the work is done, not after.  If the NSABB was concerned about this kind of research, why only express concern once the experiment succeeds?  In my intro biology class, we read another paper, published in 2005, which was addressing the exact same question, in an almost identical way.  The difference is that they failed to make a transmissible virus.  If there is a concern about this kind of research, a concern that it is too risky to do these kinds of experiments, shouldn’t the alarm have been raised regardless of the outcome?  It just doesn’t make sense to me why it suddenly became so concerning.  If anything good has come of this controversy, it is the widespread discussion that this has stimulated on the importance of open communication of scientific data, the importance of not censoring in science.  Ironically, had we all been given access to the data, like through a journal publication, it would have been apparent that there wasn’t anything to be concerned about.


Swine Flu: New and Improved!

Contributed by guest blogger: Marni Hershbain ’11

Flu season is never enjoyable, but some seasons are certainly worse than others. The 2009 swine flu outbreak was particularly serious because the 2009 H1N1 strain was a novel virus, formed via the reassortment of swine, avian and human flu viruses. There were over 600,000 confirmed cases of H1N1 and over 18,449 deaths during the course of the pandemic. While this sounds pretty bad, it could have been much worse. The transmission efficiency of H1N1 was actually much lower than those of other pandemic strains, such as the 1918 H1N1 strain. Unfortunately, recent research demonstrates that this could change.

Flu strains are characterized by the hemagglutinin and neuraminidase found on their surfaces, hence names like H1N1. In order for the virus to infect a cell, hemagglutinin on the surface of the virus must bind to glycan receptors on the cell. Therefore, to explain the low transmission efficiency of 2009 H1N1, researchers looked to its hemagglutinin.
In most flu strains, the amino acids at positions 219 and 227 within the hemagglutinin are both hydrophobic or both charged. In 1918 H1N1 both are hydrophobic. However, the 2009 H1N1 strain has isolucine, a hydrophobic molecule, in position 219 and glutamic acid, a charged molecule, in position 227. Researchers hypothesized that lacking either hydrophobic or ionic interactions at these positions would disrupt the positioning of neighboring residues and decrease the hemagglutinin’s binding affinity. They further hypothesized that if they replaced isolucine with the charged amino acid lysine, stable inter-residue interactions would occur and binding affinity would increase.

When researchers compared the ability of wild type and isolucine→lysine mutant strains to bind to an array of glycans representing human binding sites, they found the binding ability of the mutant strain was 30 times greater. The mutant version also bound more intensely to receptors in human tracheal tissue. Researchers also infected ferrets (commonly used as models in human influenza studies) with either wild type or mutant virus. Only the ferrets infected with mutant virus spread the infection to all of the previously uninfected ferrets placed in close proximity to them.

The mutation of just one amino acid could greatly impact the transmission efficiency of 2009 H1N1. Flu viruses tend to mutate frequently, which is why a new vaccine needs to be developed every year. Predicting what these mutations will be is not an easy task, but mutations at the positions in this study will certainly be monitored closely.


The Role of Social Networks in H1N1 Transmission Within a School

Contributed by Guest Blogger: Aaron Grober ’11

The H1N1 subtype of the Influenza type A virus, known colloquially as “swine flu,” was the most common cause of human influenza infection in 2009, and remained a major concern in sparking a pandemic throughout the 2009/2010 flu season.

This recent paper examines the role of grade, class, and social network in transmission of this virus in a school setting. Taking a closer look at the actual transmission pattern of this novel subtype of influenza is critical in developing models to better predict and combat pandemic spread. In the case of this school, closure due to outbreak did not significantly affect transmission among students, indicating that it may have occurred too late to be effective, stressing the importance of more exact models. The study encompassed 370 students from 295 households, surrounding an H1N1 pandemic that occurred in a Pennsylvania elementary school in April and May 2009.

The researchers found that the structuring of the school into grades and classes significantly affected the probability of transmission: 3.5% between students within a class, five times less than that between students of the same grade but different class, and five times less than that between students of different grades.

The researchers took an in-depth look at fourth-graders. They note that children are four times more likely to play with members of the same sex, and found that this behavior had a significant impact on disease transmission; the onset of epidemic transmission occurred among boys significantly before that of female classmates. In addition, they found no significant difference between recorded playmate transmission rates, and the expected proportion for if being a playmate was not a risk factor. The researchers used class seating charts to determine if proximity to an infected individual affects the risk of transmission; as it turns out, they found that sitting next to an infected individual did not significantly affect one’s risk.

In addition to school structure, the researchers looked at spread within households. The probability of a child to adult transmission within a household depended significantly on the household size, where probability of spreading the disease is much lower in larger households than smaller ones. The predominant means of adult infection was from outside the home.

These unique findings shed light on the extremely complex transmission pattern within structured populations. The biggest factors for transmission within school are grade and class, but not seating arrangement, sex, but not playmate transmission. A number of obvious questions remain: Why does sharing a class, but not a desk-space affect transmission? Why is one more likely to transmit the disease in a smaller household than a larger one? This study is an extremely insightful epidemiological tool to help explain transmission, but our knowledge of how this virus spreads remains incomplete; it seems that the flu is far more complex than we imagined.


Should I Stay or Should I Go?

Some bacteriophages (viruses that infect bacteria) can undergo a special kind of replication cycle called lysogeny. Rather than making lots of new phages upon infection, these phages can pop their genome into the host chromosome. When the cell copies its DNA and divides, the integrated phage is copied too, so the daughter cells are infected. These integrated phages, or prophages turn out to be very important to us: cholera is caused by V. cholera bacteria with an integrated phage that expresses the toxin that causes diarrhea.
So what is the evolutionary advantage of the lysogenic/lytic switch for this phage? When would it “choose” one over the other? In my biol 105 class we discussed the evolutionary benefits of lysogeny. One of the most enjoyable parts of teaching is when students ask challenging questions that lead to fruitful discussion and deeper thought. Here is my perspective on this.
You can think of the lytic cycle as highly virulent (100% mortality) or lysogenic as non-virulent (no negative effect on the host while the phage is present as a prophage). Virulence is often related to transmission, such that viruses will evolve to have the optimal level of virulence to allow for efficient transmission. Since lysogenic phages can switch between high and low virulence, when would high virulence be favored for transmission and when would low virulence be favored?
A highly virulent pathogen runs the risk of wiping out its host population. If the cells are growing actively in an environment like the gut, and the virus is replicating to high levels, it could spread to the entire host population eventually killing every cell. The virus would then depend on either more V. cholera entering the gut, or getting out of the gut and spreading to a new human host infected with cholera. Alternatively, cholera can also grow in the environment, so the phage could infect a cell in the environment. However there is some obvious risk there, that of finding the next host cholera cell either in a gut or the environment. Its a big world out there for a tiny phage and a tiny bacterial cell to meet each other. A less risky approach might be to limit virulence and allow prophage infected cells to survive. The cholera cells will be returned to the environment, where they can replicate or to a new human host where it can also replicate. Either way, the phage is guaranteed to find a host, because its already in it.
Now if the prophage finds itself in cells that are no longer growing, there may be an advantage to getting out and finding “happy” hosts. Cells that are not growing could be at greater risk of cell damage and death, perhaps they are not acquiring the nutrients and energy necessary to grow or repair cellular damage. If the cell dies, the phage will not be able to replicate. So the phage would enter the lytic cycle and release progeny. The risk of not finding a new host would presumably be lower than the risk of staying within a dying host. High virulence therefore is advantageous for transmission in this situation.
Does anyone have a different perspective?


Crop Virus Bamboozles Vectors

Contributed by Guest Blogger: H. Tran ‘14

A virus’s ability to replicate is largely dependent on the health of its host; a virus cannot proliferate in an immobile or dead organism. For this reason, viruses have a vested interest in doing as little damage as possible to ensure easy transmission. Vector-borne pathogen transmission between plants is seemingly ideal for viruses as particles can move freely from one diseased host to another potential host. However, infected plants do not typically attract vectors in the first place, as they don’t promise healthy feeding. One common crop virus is able to sidestep this obstacle by causing its diseased host to release a greater number of vector-attractants without sacrificing virulence.
It was recently discovered that the widespread plant pathogen cucumber mosaic virus elevates the release of host volatiles, or odorous chemicals, that attract vectors. Researchers measured the rates of aphid population growth on and emigration from healthy and infected plants. It was discovered that aphids were initially more attracted to the infected plants than to the healthy plants. However, the aphids dispersed rapidly from the diseased hosts after feeding. This form of transmission is known as non-persistent transmission because rather than long-term feeding and colonizing on the plants (persistent), the vectors are repelled by the inferior quality and move on to other plants (non-persistent). This type of transmission is advantageous for CMV as it facilitates easy transmission from one host to another.
It is known that the non-persistent nature of CMV transmission encourages quicker spreading between host and uninfected plants. It remains unclear, however, whether the elevated level of volatile emission is a result of an adaptation to hosts for the purpose of manipulating vectors or simply an accidental by-product of infection. In either case, the phenotypic change resulting in higher levels of volatile release has the capacity to significantly alter ecology, agriculture and human health. Damaged crops are less nutritious and unmarketable, making CMV a vastly undesirable pathogen.
Which is a more probable explanation for the deceptive mechanism by which the virus enhances host-vector interaction: manipulative adaptation or coincidentally advantageous evolution? What does your conclusion tell us about the evolution of this virus, or viruses in general? Knowing what was recently discovered about CMV, can anything be done to prevent the cultivation of diseased crops?


Social Spread of HIV

Contributed by Guest Blogger: T. McKinnon ’14

In the mid-1980’s, businesspeople were crossing the Tanzania/Uganda border, and caught a disease. This disease spread through all of Tanzania after 2 years, and this is the birth of the HIV/AIDS epidemic. The question that was being asked in this research is how rampant HIV is in two differing economic classes, the “rich” and the “poor,” and in which is it more prevalent. The model created by studying the transmission of this disease through differing socio-economic classes is to see the impact HIV/AIDS has on one economic class versus another, and whether transmission is easier, harder, faster, etc. in different social classes.
The experiment conducted worked like this: a total population of individual is accounted for, divided into susceptibles, infectives (infectious), pre-AIDS and AIDS patients. These people are then divided into pre-AIDS hospitalized patients and AIDS patients seeking no hospitalization, because this is common in lower economic classes. From then, the spread and rate of infection of HIV and the spread of AIDS is measured among these separate groups, whether it is initial infection or development into full blown AIDS.
Through extensive experimentation, HIV/AIDS was found to be more prevalent among wealthier populations, but it spreads faster among the lower classes. I find it very interesting that this disease is not more prevalent and spreads faster in the lower classes. In the upper class, people can more readily afford the treatments and medications than people living in lower classes with less money.
The researchers acknowledged that this experiment was by no means exhaustive. I would like this experiment to expand to how race and sexuality interact with social class in the spread and prevalence of HIV/AIDS or if race has anything to do with it, both separately and together. I also would like to know how level of sexual activity among social class propagates HIV spread, and if the members of the upper class were more or less sexually active, or participated in more unsafe sexual practices than those of the lower class, or if it was the other way around.


Cross-Species Transmission in Rabies

Contributed by Guest Blogger: D. Patel ’14

Deadly human diseases including HIV Aids, swine flu and rabies are infectious diseases where the viruses have jumped from one animal species into another and now infect humans too. This is a phenomenon known as cross-species transmission (CST). Understanding this process is the key to predicting and preventing future outbreaks.
The scientists who researched CST and wrote this paper made a groundbreaking discovery into how viruses jump from host to host. They used and thought of rabies as an ideal system because it occurs across the country, affects many different host species, and is known to mutate frequently. Although cases of rabies in humans are rare in the U.S., bats are a common source of infection. Hence, the study was based on and narrowed down to CST events among different bat species.
To determine the rate of CST, a large dataset containing hundreds of rabies viruses from 23 North American bat species was used. Population genetics tools were used to quantify how many CST events were expected to occur from any infected individual and the cases were verified by genotyping both the viruses and the bats.
The study showed that depending on the species involved, a single infected bat may infect between 0 and 1.9 members of a different species; and that, on average, CST occurs only once for every 72.8 transmissions within the same species. This means that the majority of viruses from cross-species infections were tightly nested among genetically similar bat species.
It is a long-held belief that CST depends on virus mutation and contact of the host with other species. However, this study showed that CST may have more to do with host similarity. The similarity in the defenses of closely related species may favor virus exchange by making it easier for natural selection to favor a virus’ ability to infect new hosts.
Whether other factors (like evolution of viruses) are enough to overcome the genetic differences between hosts remains questionable. However, the basic knowledge gained through the study is key to developing new intervention strategies for diseases that can jump from wildlife to humans.