Archive for the ‘Uncategorized’ Category

The Assembly Pathway of an Icosahedral Single-Stranded RNA Virus Depends on the Strength of Inter-Subunit Attractions

27 March, 2014

See on Scoop.itVirology News

The strength of attraction between capsid proteins (CPs) of cowpea chlorotic mottle virus (CCMV) is controlled by the solution pH. Additionally, the strength of attraction between CP and the single-stranded RNA viral genome is controlled by ionic strength. By exploiting these properties, we are able to control and monitor the in vitro co-assembly of CCMV CP and single-stranded RNA as a function of the strength of CP–CP and CP–RNA attractions. Using the techniques of velocity sedimentation and electron microscopy, we find that the successful assembly of nuclease-resistant virus-like particles (VLPs) depends delicately on the strength of CP–CP attraction relative to CP–RNA attraction. If the attractions are too weak, the capsid cannot form; if they are too strong, the assembly suffers from kinetic traps. Separating the process into two steps—by first turning on CP–RNA attraction and then turning on CP–CP attraction—allows for the assembly of well-formed VLPs under a wide range of attraction strengths. These observations establish a protocol for the efficient in vitro assembly of CCMV VLPs and suggest potential strategies that the virus may employ in vivo.


Ed Rybicki‘s insight:

I do love it when a paper is published that could have been done pretty much any time in the last 40 years – and with one of my favourite viruses, that I played with a LOT back there before 1980.

Ultracentrifugation, pH meters, ionic strength determinations, EM…all tried and true, and used 40+ years ago. OK, they also used cloned BMV RNA 1 cDNA, and did 3-D image reconstruction from EMs, but hey, they needn’t have done that!  Nice, straightforward physicochemical studies, on a well-characterised virus, with good, simple conclusions. 

Namely, that assembly of the virus is NOT just a simple mix-CP-and-RNA-and-it-will-happen, but that it depends upon both pH, for modulating ionic interactions,and ionic strength for modulating ionic interactions AND the "hydrophobic effect", as we used to know it.

While their conclusions are relevant for assembly of heat- and nuclease-resistant nano particles in vitro, I wonder if they are physiologically relevant: if "correct" assembly depends upon first, turning on CP-RNA attraction (ionic strength shift), and second, turning on CP-CP attraction (pH shift) – where in the cell does that happen?

In their own words, "It is generally accepted that the cytoplasm of plant cells is maintained near neutral pH with ionic strength of approximately 0.1 M. Our in vitro results show that these conditions are insufficient for nucleocapsid formation in the absence of cellular host factors."

Yeeee-ee-eesssss…precisely. What happens in the cell? The answer could lie in the one thing they don’t report, but that some of the heroes of my distant youth – people like JB Bancroft and Thom Hohn, both quoted (from 1970 and 1969 respectively) in this paper, DID do. Namely, investigate what happens at different CP and RNA concentrations, at constant pH and ionic strength.

You see, it was shown 30+ years ago – and I have been lecturing on it since then – that CP and RNA for viruses like BMV / CCMV and MS2 form different complexes with their cognate partners at different molecular ratios. That is, at low CP:RNA ratios, a high-affinity complex is formed, which is basically a ribonucleoprotein complex without structure. Increasing the CP:RNA ratio for both MS2 and CCMV, as I recall (maybe Dick Verduin was involved with CCMV), results in further lower-affinity association of CP with both RNA and already-bound CP – which acts as a nucleation complex – to result in full capsid assembly.

I note that the process in both cases was shown to be specific, for low CP:RNA ratios: that is, it was cognate CP and RNA doing the high affinity nucleation complex formation.

And these guys deliberately used a heterologous RNA…albeit one from a related virus, but still: what would have happened if they’d used CCMV RNA?

Still - great paper, taking me back to when I wrote an essay on "Assembly of Spherical Plant Viruses" in my Honours year in 1977, quoting quite a few of the same references these folk did. Ah, simpler times…B-)

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This Old PI….

7 March, 2014

See on Scoop.itVirology News

Ed Rybicki‘s insight:

I blame Chris Upton

GM crops: European scientists descend on Africa to promote biotech

24 February, 2014

See on Scoop.itVirology News

Africa is expected to be the next target of GM food companies, as European scientists and policymakers travel to Ethiopia to boost the prospect of growing more of the controversial crops on the continent.

Anne Glover, the chief scientific adviser to the European commission, and other prominent pro-GM researchers and policymakers from European countries including Germany, Hungary, Italy and Sweden will this week meet Ethiopian, Kenyan, Ghanaian and Nigerian farm ministers as well as officials from the African Union.

Ed Rybicki‘s insight:

And that’ll go down REALLY well: European governments and policymakers pushing GM crops in Africa, when their own people don’t want them?  

Whether those reasons are stupid or not, and whether or not it is a good idea to plant GM in Africa, I can see a knee-jerk anti-Europe response could well work to completely screw up just what it is they are trying to do.

Consider: Zimbabwe and Zambia resolutely oppose even the import of GM maize as FOOD, let alone allowing the planting of it.  Africans have a history of being VERY suspicious of outsiders bearing gifts – because there is a history of dumping stuff in Africa, of everything from suspect pharmaceuticals to excess chickens.

I predict a lead balloon result for this conference and for the initiative.  Time for Europe to listen to the Kenyans, Burkinabe and South Africans about the merits of growing GM – and not the other way around!

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South Africa unable to produce foot-and-mouth disease virus vaccine

20 February, 2014

See on Scoop.itVirology News

Outbreak of foot and mouth disease four years ago largely due to collapse of state’s ability to manufacture vaccines

THE outbreak of foot and mouth disease four years ago, which cost South Africa R4bn a year in lost exports, was largely due to the collapse of the state’s ability to manufacture vaccines.

Agriculture, Forestry and Fisheries Minister Tina Joemat-Pettersson told a press briefing on Wednesday on the lifting the ban of red meat exports by the World Animal Health Organisation (WAHO). The removal of the boycott will not affect red meat producers significantly as South Africa usually imports the product to satisfy domestic demand. Only about 1% of red meat production, mainly venison, is exported. Other animal exports include hides and wool.

Ed Rybicki‘s insight:

This is actually horrifying, in the context of what is SUPPOSED to be Africa’s most developed economy – and especially its most developed agricultural economy.

Really: it is a fact not much appreciated outside of the agricultural sector, that we have to get our FMDV vaccines from BOTSWANA.

That’s right – the country with the largest economy in Africa gets its FMDV vaccines from the Botswana Veterinary Institute, a factory set up by the French in a country with a total population about a quarter of that of Greater Johannesburg.

So, let us get this straight: the country that owns the Onderstepoort Veterinary Institute, where the legendary Sir Arnold Theiler pioneered virus and other vaccines from the 1890s on, cannot make its own FMDV vaccines any more?

The country that USED to make a whole range of animal vaccines via the state-owned facility that is now Onderstepoort Biological Products, is now struggling to make just a few?

The country that once built a state-of-the-art BSL4 facility dedicated to FMDV now cannot operate it, or even make old-style killed vaccines?

Don’t let me get started on human vaccines, because we now don’t make ANY – but the erosion of our capacity to make vaccines means our ability to look after human and animal health is now severely under threat.

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RNA sequencing of 750-year-old barley virus sheds new light on the Crusades

7 February, 2014

See on Scoop.itVirology News

Scientists have for the first time sequenced an ancient RNA genome – of a barley virus once believed to be only 150 years old – pushing its origin back at least 2,000 years and revealing how intense farming at the time of the Crusades contributed to its spread.

Ed Rybicki‘s insight:

This is a good report about a VERY interesting finding – with one flaw.  They go on, apparently, about how BSMV is a "new virus": why would anyone think that?  Since the late 1980s, Adrian Gibbs and others have pointed out that tobamoviruses are probably ancient; just because RNA CAN evolve fast doesn’t mean it does, in terms of encoding functional elements.  Gibbs showed this for plant viruses; it has also been done for the HIV/SIV complex, where it is shown that a similar divergence in sequence among theses viruses to all animals since the Cretaceous, has led to NO changes in morphology, or gene function.

The simple fact is that having "plastic" genome in comparison to eukaryotic cells does NOT mean that ssRNA viruses may not be ancient.

Having said all that, it really is a tour de force to have sequenced a virus that old – aided by the fact, I am sure, that BSMV is hardy little beast, with a really stable vision.

Now, to find those hundred-year-old maize leaves put away with maize streak virus symptoms….B-)

Thanks to @Elsevier Microbiol* for pointing this out!

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Proof of principle for epitope-focused vaccine design – or is it?

6 February, 2014

See on Scoop.itVirology News

Vaccines prevent infectious disease largely by inducing protective neutralizing antibodies against vulnerable epitopes. Several major pathogens have resisted traditional vaccine development, although vulnerable epitopes targeted by neutralizing antibodies have been identified for several such cases. Hence, new vaccine design methods to induce epitope-specific neutralizing antibodies are needed. Here we show, with a neutralization epitope from respiratory syncytial virus, that computational protein design can generate small, thermally and conformationally stable protein scaffolds that accurately mimic the viral epitope structure and induce potent neutralizing antibodies. These scaffolds represent promising leads for the research and development of a human respiratory syncytial virus vaccine needed to protect infants, young children and the elderly. More generally, the results provide proof of principle for epitope-focused and scaffold-based vaccine design, and encourage the evaluation and further development of these strategies for a variety of other vaccine targets, including antigenically highly variable pathogens such as human immunodeficiency virus and influenza.

Ed Rybicki‘s insight:

Yes.  Um.  Well.  I have just heard a lecture by the redoubtable Marc van Regenmortel (disclosure: my former mentor) pointing out how this strategy is highly UNLIKELY to be "the" way of generating antigens that are likely to be good immunogens – and I was convinced.

Now this.

Um, he said.  And…the MHvR arguments still stand, I think: why should something that supposedly looks like the thing that supposedly elicited a strongly neutralising antibody, elicit a strongly neutralising antibody?

OK, it apparently did here – but as a general rule?  Do we KNOW what "good" neutralising epitopes look like in 3D?  Because linear epitopes are really just an approximation to what real epitopes are, and epitopes are in fact just those things that BIND antibodies – NOT necessarily what ELICITS them.

Damn, I’m sounding like Marc – must be catching!

But seriously: antibodies are the membrane receptors of B cells; they are generated by random recombinations, and then selected by adventitiously binding another molecule that is not self; they are then further selected by repeated rounds of binding, while undergoing random mutations affecting their hypervariable regions, while weak binders are selected AGAINST by Tregs.

And end up both with higher affinity, and more broad binding to related "epitopes" as a result.

Incredibly hard to duplicate as general phenomenon, I would surmise.

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Have we reached the beginning of the end of the fight against HIV/AIDS?

5 February, 2014

See on Scoop.itVirology News

The world is at a tipping point in the fight against HIV/Aids with the number of people getting treatment set to be greater than those being newly diagnosed. But have we really reached the beginning of the end?

Considering the first World Aids Day took place 25 years ago in 1988, it’s a remarkable achievement. Today, there are about 35million people with HIV and Aids in the world, with 9.7million receiving treatment that can enable a long and healthy life.

Ten years ago, just 300,000 people received such treatment so it’s clear there have been major gains, and although experts, campaigners and health practitioners are buoyed the ‘tipping point’ is on the horizon, hard work lies ahead. Erin Hohlfelder, global health policy director of the One Foundation and author of its report The Beginning Of The End?’, feels reaching the tipping point will be a landmark in the fight against HIV and Aids. ‘The disease has been outpacing us for decades, with more new infections than people being added to treatment.

‘To use an analogy, if you had five cuts on your hand, we’ve only had one plaster to treat the cuts, so you’re still bleeding,’ she said.

‘The tipping point, where we have more people living healthy lives through treatment for HIV, than newly infected people, means we’ve caught up with the disease and begun to get ahead of it, so it’s a critical moment. It’s a marker that signals for the first time HIV and Aids is on a downward curve,’ said Ms Hohlfelder.

Ed Rybicki‘s insight:

Great infographic – and potentially good news.  But as I have told my students repeatedly since the mid-1980s, this is a long. slow pandemic – and we are only just over halfway through it.

There’s still a long way to go.

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Cases of New Deadly Bird Flu Surge in China, Experts Say

5 February, 2014

See on Scoop.itVirology News

About 300 cases of H7N9 avian influenza have been confirmed in China, with more appearing every day.

Cases of the new H7N9 avian influenza in China are surging alarmingly, flu experts warned this week.

There are now about 300 confirmed cases, with more appearing every day. Roughly a quarter of the victims have died.

The first human cases were reported only last March. By contrast, the H5N1 influenza virus, another lethal strain that jumped from birds to people, first appeared in 2003 and took almost five years to reach the 300-case mark.

Ed Rybicki‘s insight:

So THIS is the next one?  Possibly?  Good job they’re <a href="">already making vaccines</a>, then, isn’t it?


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Biodiversity: So much more than legs and leaves

28 January, 2014

See on Scoop.itAquatic Viruses

Microorganisms inhabit virtually every possible niche on Earth, including those at the outer envelope of survival. However, the focus of most conservation authorities and ecologists is the ‘legs and leaves’ side of biology -the ‘macrobiology’ that can be seen with the naked eye. There is little apparent concern for the preservation of microbial diversity, or of unique microbial habitats. Here we show examples of the astounding microbial diversity supported by South Africa’s ecosystems and argue that because microbes constitute the vast majority of our planet’s species they should be considered seriously in the future protection of our genetic resources.


Ed Rybicki‘s insight:

This is our stab at getting South African science funders to take microbial diversity seriously.  I helped write it, obviously.  And it’s Open Access!

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RNA’s Secret Life Outside the Cell – Wired Science

21 January, 2014

See on Scoop.itVirology News

Circulating RNAs carry messages between the cells of plants and invertebrates. Do they do the same for us?

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