TWiV 411: Chicken runs

The TWiVeroos examine a reverse spillover of Newcastle disease virus vaccines into wild birds, and identification of a protein cell receptor for murine noroviruses.

You can find TWiV #411 at, or listen below.

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Eukaryotic genes in a bacteriophage


Wolbachia in an insect cell. Image credit: PLoS/Scott O’Neill.

Viruses are tidily categorized into three groups according to the hosts they infect – bacteriophages, eukaryotic viruses, and archaeal viruses. Viruses do not infect hosts in another domain of life, and therefore lateral gene transfer is limited (giant DNA viruses might be exceptions). Now there is evidence for lateral gene transfer between eukaryotes and bacteriophages.

Proof of this unusual movement of DNA comes from studies of the obligate intracellular bacteria Wolbachia, which infects 40% of arthropods (pictured). Wolbachia are in turn infected with a bacteriophage called WO; nearly all sequenced Wolbachia genomes contain integrated WO DNA. Analysis of complete WO genome sequences revealed the presence of mutiple eukaryotic genes (link to paper) that comprise about half of the phage genome!

Ten different protein domains were identified in the eukaryotic genes of WO phage with four functions: toxins, host-microbe interactions, host cell suicide, and protein secretion through membranes.

One eukaryotic gene in phage WO is a black widow spider toxin called latrotoxin-CTD. Sequence analysis suggests that the spider toxin gene was transferred to phage WO within a Wolbachia genome (these bacteria are known to infect widow spiders).

It is not surprising that a virus of a bacterium that infects a eukaryotic cell might acquire eukaryotic genes, but the exact mechanism of gene transfer is unknown. Eukaryotic DNA might enter the WO genome while the particles are in the insect cell cytoplasm, or during packaging of viral DNA in the presence of animal DNA. Another possibility is transfer of eukaryotic DNA to the Wolbachia genome, and then to phage WO.

The fact that eukaryotic-like DNA sequences make up half of the phage WO genome suggests that they serve important functions for the virus. The functions ascribed to these eukaryotic genes suggest roles in cell lysis, modification of host proteins, and toxicity.

There are other examples of phage-infected obligate intracellular bacteria of Chlamydia, aphids, and tsetse flies. A study of these viral genomes should reveal whether lateral gene transfer between metazoans and bacteriophages is a common mechansim for augmenting functions of the viral genome.

Virus Watch: Cancer Killing Viruses

Guest host Lynda Coughlan reviews how oncolytic viruses, which specifically kill tumor cells, are designed and how they work.

TWiV 410: Hurricane Zika

Sharon and Scott join the TWiV team to talk about their work on dengue antibody-dependent enhancement of Zika virus infection, and identifying the virus in mosquitoes from Miami.

You can find TWiV #410 at, or listen below.

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A Nobel for autophagy, and the importance of fundamental research

autophagyYoshinori Ohsumi has been awarded the 2016 Nobel Prize for Physiology or Medicine for his work on autophagy, a process of eukaryotic cells for degrading and recycling cellular components. Because of his research, we now understand the importance of autophagy in health and human disease. It is another example of the serendipity of science and yes, it is relevant to virology!

The word autophagy was coined by Christian de Duve in 1963 to describe a process that he and others had previously described: when stressed, cells would sequester portions of the cytoplasm in double-membraned vesicles called autophagosomes. These would then fuse with lysosomes (which de Duve had discovered) and the contents were degraded (illustrated; image credit).

In subsequent years it was suggested that autophagy might have roles in human disease, but little progress was made on understanding how the process worked: how it was triggered, what proteins were involved, and its function in health and pathogenesis.

As a young Assistant Professor at Tokyo University in the early 1990s, Ohsumi found that autophagy occurred in the yeast Saccharomyces cerevisiae. He decided to produce yeast strains lacking the proteases that would be involved in digesting the contents of the autophagic vesicles, with the idea that these vesicles would accumulate under stress (under normal conditions the autophagosome exists for a short period of time, making its study difficult).

When Ohsumi stressed the yeasts lacking the vacuolar (equivalent of mammalian lysosome) proteases, autophagosomes accumulated in the cytoplasm which were readily visible by light microscopy. He used this clear phenotype to isolate yeast mutants that could no longer accumulate such vesicles, and identified 15 genes that are essential for induction of autophagy in eukaryotic cells.

Later Ohsumi elucidated the functions of these genes in autophagy induction, and found homologues in mammalian cells. His work stimulated great interest in autophagy, and it became a highly studied pathway. We now understand that autophagy is not only important for embryogenesis and cell differentiation, but plays roles in neurodegenerative diseases, cancer, and in defenses against bacterial and viral infections.

As might be expected, virus infection is a stress that triggers autophagy, which may impact the outcome of infections: it can have both antiviral functions, and it can also stimulate virus replication. As might be expected for a cellular process that degrades cytosolic contents, autophagy can lead to clearance of viruses. Consequently a number of viral genomes encode proteins that inhibit autophagy, including herpes simplex virus. Degradation of viral proteins by autophagy can also provide peptides for presentation to the cellular adaptive immune system, further enhancing clearance.

Autophagy may also benefit virus replication. For example, non-lytic cell to cell spread of poliovirus depends on release of virus particles from autophagosomes, and autophagy of lipids provides metabolic energy for dengue virus replication.

When autophagy was first described in the 1950s, no one knew its significance. Nevertheless, a number of scientists, including Ohsumi, continued to study autophagy because they were curious. The unexpected result was the elucidation of a pathway that has substantial roles in a variety of human diseases.

The lesson is clear: let scientists pursue their curiosity. It’s fine to target specific research problems, like curing cancer or diabetes, but don’t ignore fundamental research on problems that don’t seem to be directly relevant to human diseases. I’m concerned (as are many others) that the US science establishment is moving away from fundamental research (whose benefits may not be apparent for a long time) to translational research.

How do we explain the trend from fundamental to translational research? I don’t have all the answers, but I think part of the problem is that the US Congress likes to spend taxpayer money on targeted problems, like Alzheimer’s disease. But tell them that you want to study a process in cells that looks interesting, but you are not sure what it means, and you can see their reluctance to support this type of work.

TWiV 409: A Nef is enough

Jeremy joins the TWiVeroids to tell the amazing story of how the function of the HIV-1 protein called Nef was discovered and found to promote infection by excluding the host protein SERINC from virus particles.

You can find TWiV #409 at, or listen below.

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Three countries endemic for poliovirus

poliovirusI cannot let September pass without noting that 34 years ago this month, I arrived at Columbia University to start my laboratory to do research on poliovirus (pictured). That virus is no longer the sole object of our attention – we are wrapping up some work on poliovirus and our attention has shifted elsewhere. But this is a good month to think about the status of the poliovirus eradication effort.

So far this year 26 cases of poliomyelitis have been recorded – 23 caused by wild type virus, and three caused by vaccine-derived virus. At the same time in 2015 there were 44 reported cases of polio – small progress, but, in the words of Bill Gates, the last one percent is the hardest.

One of the disappointments this year is Nigeria. It was on the verge of being polio-free for one year – the last case of type 1 poliovirus in Nigeria had been recorded in July of 2014. In August the government reported that 2 children developed polio in the Borno State. The genome sequence of the virus revealed that it had been circulating undetected in this region since 2011. Due to threats from militant extremists, it has not been possible for vaccination teams to properly cover this area, and surveillance for polioviruses has also been inefficient. The virus can circulate freely in a poorly immunized population, and as only 1% of infections lead to paralysis, cases of polio might have been missed.

The conclusion from this incident is that the declaration that poliovirus is no longer present in any region is only as good as the surveillance for the virus, which can never be perfect as all sources of infection cannot be covered.

Of the 26 cases of polio recorded so far in 2016, most have been in Afghanistan and Pakistan (9 and 14, respectively). It is quite clear that conflict has prevented vaccination teams from immunizing the population: in Pakistan, militants have attacked polio teams during vaccination campaigns.

Recently 5 of 27 sewage samples taken from different parts of the province of Balochistan in Pakistan have tested positive for poliovirus. Nucleotide sequence analysis revealed that the viruses originated in Afghanistan. The fact that such viruses are present in sewage means that there are still individuals without intestinal immunity to poliovirus in these regions. In response to this finding, a massive polio immunization campaign was planned for the end of September in Pakistan. This effort would involve 6000 teams to reach 2.4 million children. Apparently police will be deployed to protect immunization teams (source: ProMedMail).

The success of the polio eradication program so far has made it clear that if vaccines can be deployed, circulation of the virus can be curtailed. If immunization could proceed unfettered, I suspect the virus would be gone in five years. But can anyone predict whether it will be possible to curtail the violence in Pakistan, Afghanistan, and Nigeria that has limited polio vaccination efforts?

TWiV 408: Boston Quammens

Four years after filming ‘Threading the NEIDL’, Vincent and Alan return to the National Emerging Infectious Diseases Laboratory BSL4 facility at Boston University where they speak with science writer David Quammen.

You can find TWiV #408 at, or watch/listen here.

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TWiV 407: Tar Heels go viral, part one

In the first of two shows recorded at the University of North Carolina in Chapel Hill, Vincent meets up with faculty members to talk about how they got into science, their research on DNA viruses, and what they would be doing if they were not scientists.

You can find TWiV #407 part one at Or watch the video above, or listen below.

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Structure of an infectious prion

prion conversionPrions are not viruses – they are infectious proteins that lack nucleic acids. Nevertheless, virologists have always been fascinated by prions – they appear in virology textbooks (where else would you put them?) and are taught in virology classes. I’ve written about prions on this blog (five articles, to be exact – look under P in the Table of Contents) and I’m fascinated by their biology and transmission. That’s why the newly solved structure of an infectious prion protein is the topic of the sixth prion article at virology blog.

Spongiform encephalopathies are neurodegenerative diseases caused by misfolding of normal cellular prion proteins. Human spongiform encephalopathies are placed into three groups: infectious, familial or genetic, and sporadic, distinguished by how the disease is acquired initially. In all cases, the pathogenic protein is the host-encoded PrPC protein with an altered conformation, called PrPsc. In the simplest case, PrPSc converts normal PrPC protein into more copies of the pathogenic form (illustrated).

The structure of the normal PrPprotein, solved some time ago, revealed that it is largely alpha-helical with little beta-strand content. The structure of PrPSc protein has been elusive, because it forms aggregates and amyloid fibrils. It has been suggested that the PrPSc protein has more beta-strand content than the normal protein, but how this property would lead to prion replication was unknown. Clearly solving the structure of prion protein was needed to fully understand the biology of this unusual pathogen.

The structure of PrPSc protein has now been solved by cryo-electron microscopy and image reconstruction (link to paper). The protein was purified from transgenic mice programmed to produce a form of  PrPSc protein that is not anchored to the cell membrane, and which is also underglycosylated. The protein causes disease in mice but is more homogeneous and forms fibrillar plaques, allowing gentler purification methods.

prion structureThe structure of this form of the PrPSc protein reveals that it consists of two intertwined fibrils (red in the image) which most likely consist of a series of repeated beta-strands, or rungs, called a beta-solenoid. The structure provides clues about how a pathogenic prion protein converts a normal PrPC into PrPSc . The upper and lower rungs of beta-solenoids are likely the initiation points for hydrogen-bonding with new PrPC molecules – in many proteins with beta-solenoids, they are blocked to prevent propagation of beta-sheets. Once added to the fibrils, the ends would serve to recruit additional proteins, and the chain lengthens.

The authors note that the molecular interactions that control prion templating, including hydrogen-bonding, charge and hydrophobic interactions, aromatic stacking, and steric constraints, also play roles in DNA replication.

The structure of PrPSc protein provides a mechanism for prion replication by incorporation of additional molecules into a growing beta-solenoid. I wonder if incorporation into fibrils is the sole driving force for converting PrPCprotein into PrPSc, or if PrPC is conformationally altered before it ever encounters a growing fibril.