What Would Happen if You Got Ebola?

13 August, 2014

A secondary infection in the U.S. is highly unlikely. But here’s how the healthcare system would respond if there was one.

Source: www.theatlantic.com

Goes without saying that this would happen in a lot of other places, too.  Including our very own South Africa – where it HAS happened, with Marburg, Ebola and Lujo viruses.  Written about right here on ViroBlogy.

See on Scoop.itVirology News

How can geminiviral Rep capture the cell cycle of differentiated plant cells?

12 August, 2014

African cassava mosaic virus (ACMV) in the geminivirus family has being affected 500 million people worldwide by devastating cassava crops during the past decades. It has caused severe symptoms and reduced yield up to the complete loss of roots, the main starchy food source especially for subsistence farmers in Africa. How can a tiny virus with a small genome evoke such dramatic effects? The viral key component, the replication-initiator protein (Rep), forces differentiated plant cells in the phloem to reactivate DNA synthesis. Even more, it does the same in model cells of fission yeast. We have identified, now, a potential cyclin interaction motif, RXL, in the sequence of ACMV Rep, which may be important for cell cycle control. This motif is essential to induce rereplication in yeast and necessary for viral infection of plants.

 

Source: www.virologyhighlights.com

I am a sucker for geminiviruses and their replication – as can be seen in the pages published here and elsewhere over the years.  It is fascinating to me that a small protein like Rep – only ~30 kDa – can do so many things, and especially interfere in such a fundamental way with organised, differentiated cells.

What is even more interesting is that it can do it in such a wide variety of systems: it’s been shown that ACMV can replicate in maize protoplasts as well as in the dicotyledonous cassava; it can evidently function well in yeast as well – and via a pathway that no-one suspected before now.

Truly, a protein of many parts!  Congratulations to Katharina Hipp and to my old friends Bruno and Holger.

See on Scoop.itVirology News

Plant-made antibodies used as therapy for Ebola in humans: post-exposure prophylaxis goes green!

5 August, 2014
Ebola virus budding from an infected cell.  Courtesy of Russell Kightley Media

Ebola virus budding from an infected cell.
Courtesy of Russell Kightley Media

Yes, I know you fans of ViroBlogy like Ebola – and just coincidentally, I was desperately trying to finish a review* on “Plant-based vaccines against viruses” against a backdrop of an out-of-control Ebola epidemic in West Africa, when three different people emailed me different links to news of use of a plant-made monoclonal antibody cocktail.  I immediately included it in my review – and I am publishing an excerpt here, for informations’ sake.  Enjoy!

Plantibodies against Ebola

The production of anti-Ebola virus antibodies has recently been explored in plants: this could yet become an important part of the arsenal to prevent disease in healthcare workers, given that at the time of writing an uncontrolled Ebola haemorrhagic fever outbreak was still raging in West Africa, and the use of experimental solutions was being suggested (Senthilingam, 2014). For example, use of a high-yielding geminivirus-based transient expression system in N benthamiana that is particularly suited to simultaneous expression of several proteins allowed expression of a MAb (6DB) known to protect animals from Ebola virus infection, at levels of 0.5 g/kg biomass (Chen et al., 2011). The same group also used the same vector system (described in detail here (Rybicki and Martin, 2014)) in lettuce to produce potentially therapeutic MAbs against both Ebola and West Nile viruses (Lai et al., 2012).

A more comprehensive investigation was reported recently, of both plant production of Mabs and post-exposure prophylaxis of Ebola virus infection in rhesus macaques (Olinger et al., 2012). Three Ebola-specific mouse-human chimaeric MAbs (h-13F6, c13C6, and c6D8; the latter two both neutralising) were produced in whole N benthamiana plants via agroinfilration of magnICON TMV-derived viral vectors. A mixture of the three MAbs – called MB-003 – given as a single dose of 16.7 mg/kg per Mab 1 hour post-infection followed by doses on days 4 and 8, protected 3 of 3 macaques from lethal challenge with 1 000 pfu of Ebola virus. The researchers subsequently showed significant protection with MB-003 treatment given 24 or 48 hours post-infection, with four of six monkeys testing surviving, compared to none in two controls. All surviving animals treated with MB-003 experienced insignificant if any viraemia, and negligible clinical symptoms compared to the control animals. A significant finding was that the plant-produced MAbs were three times as potent as the CHO cell-produced equivalents – a clear case of plant production leading to “biobetters”. A follow-up of this work investigated efficacy of treatment with MB-003 after confirmation of infection in rhesus macaques, “according to a diagnostic protocol for U.S. Food and Drug Administration Emergency Use Authorization” (Pettitt et al., 2013). In this experiment 43% of treated animals survived, whereas all controls tested here and previously with the same challenge protocol died from the infection.

In news from just prior to submission of this article, a report quoted as coming from the National Institute of Allergy and Infectious Diseases states that two US healthcare workers who contracted Ebola in Liberia were treated with a cocktail of anti-Ebola Mabs called ZMapp – described as a successor to MB-003 – developed by Mapp Pharmaceutical of San Diego, and manufactured by Kentucky BioProcessing (Langreth et al., 2014). Despite being given up to nine days post-infection in one case, it appears to have been effective (Wilson and Dellorto, 2014).

A novel application of the same technology was also used to produce an Ebola immune complex (EIC) in N benthamiana, consisting of the Ebola envelope glycoprotein GP1 fused to the C-terminus of the heavy chain of the humanised 6D8 MAb, which binds a linear epitope on GP1. Geminivirus vector-mediated co-expression of the GP1-HC fusion and the 6D8 light chain produced assembled immunoglobulin, which was purified by protein G affinity chromatography. The resultant molecules bound the complement factor C1q, indicating immune complex formation. Subcutaneous immunisation of mice with purified EIC elicited high level anti-GP1 antibody production, comparable to use of GP1 VLPs (Phoolcharoen et al., 2011). This is the first published account of an Ebola virus candidate vaccine to be produced in plants.

References

Chen, Q., He, J., Phoolcharoen, W., Mason, H.S., 2011. Geminiviral vectors based on bean yellow dwarf virus for production of vaccine antigens and monoclonal antibodies in plants. Human vaccines 7, 331-338.

Lai, H., He, J., Engle, M., Diamond, M.S., Chen, Q., 2012. Robust production of virus-like particles and monoclonal antibodies with geminiviral replicon vectors in lettuce. Plant biotechnology journal 10, 95-104.

Langreth, R., Chen, C., Nash, J., Lauerman, J., 2014. Ebola Drug Made From Tobacco Plant Saves U.S. Aid Workers. Bloomberg.com.

Olinger, G.G., Jr., Pettitt, J., Kim, D., Working, C., Bohorov, O., Bratcher, B., Hiatt, E., Hume, S.D., Johnson, A.K., Morton, J., Pauly, M., Whaley, K.J., Lear, C.M., Biggins, J.E., Scully, C., Hensley, L., Zeitlin, L., 2012. Delayed treatment of Ebola virus infection with plant-derived monoclonal antibodies provides protection in rhesus macaques. Proceedings of the National Academy of Sciences of the United States of America 109, 18030-18035.

Pettitt, J., Zeitlin, L., Kim do, H., Working, C., Johnson, J.C., Bohorov, O., Bratcher, B., Hiatt, E., Hume, S.D., Johnson, A.K., Morton, J., Pauly, M.H., Whaley, K.J., Ingram, M.F., Zovanyi, A., Heinrich, M., Piper, A., Zelko, J., Olinger, G.G., 2013. Therapeutic intervention of Ebola virus infection in rhesus macaques with the MB-003 monoclonal antibody cocktail. Science translational medicine 5, 199ra113.

Phoolcharoen, W., Bhoo, S.H., Lai, H., Ma, J., Arntzen, C.J., Chen, Q., Mason, H.S., 2011. Expression of an immunogenic Ebola immune complex in Nicotiana benthamiana. Plant biotechnology journal 9, 807-816.

Rybicki, E.P., Martin, D.P., 2014. Virus-Derived ssDNA Vectors for the Expression of Foreign Proteins in Plants. Current topics in microbiology and immunology 375, 19-45.

Senthilingam, M., 2014. Ebola outbreak: Is it time to test experimental vaccines? CNN.

Wilson, J., Dellorto, D., 2014. 9 questions about this new Ebola drug. CNN.

* = which, despite their having commissioned from me, the good folk at “Viruses” an unnamed journal decided “…may not have substantial differences with the reviews you published recently” – and rejected.  I shall have revenge.  Oh, yes…B-)

2013 in review: ViroBlogy seems to be doing OK??

5 August, 2014

The WordPress.com stats helper monkeys prepared a 2013 annual report for this blog.

Here’s an excerpt:

The concert hall at the Sydney Opera House holds 2,700 people. This blog was viewed about 24,000 times in 2013. If it were a concert at Sydney Opera House, it would take about 9 sold-out performances for that many people to see it.

Click here to see the complete report.

Has the Time Come to Test Experimental Ebola Vaccines?

30 July, 2014

The Ebola Virus is Spreading Across West Africa in The Largest Outbreak To Date

Mortality rates are currently at 60%, where normally up to 90% of affected people die. Unfortunately, there are no cures or vaccines for the disease, despite its emergence in 1976.

In March of this year, cases of Ebola were reported for the first time in Guinea, Western Africa. Notably, these outbreaks took place in Guinean districts that bordered Sierra Leone and Liberia. In the past, outbreaks have taken place in remote areas of Africa, but this outbreak has had the opportunity to cross border and spread throughout Western Africa, and it has done just that. Now fourth months since the March outbreak, 1,093 people have been infected and there have been a reported 660 deaths attributed to the deadly virus (source: CNN vital signs.)

 

Source: www.vaccinenation.org

I would say – yes!  I am sure the beleaguered healthcare workers in Guinea, Liberia, Sierra Leone and now Nigeria would welcome the experimental vaccine candidates, and the experimental therapeutics, for that matter.

See on Scoop.itVirology News

“Controversial scientist recreates H1N1 flu that killed 500K people” – NOT

2 July, 2014

Dr Yoshihiro Kawaoka, professor of virology at University of Wisconsin at Madison, has tweaked the 2009 strain of pandemic influenza to make it resistant the human immune system’s antibodies.

Source: www.dailymail.co.uk

Trust the Dimwitted Mail to misstate what happened – which is that Yoshihiro Kawaoka selected the H1N1pdm 2009 flu virus in culture till he came up with antibody-binding escape mutants.

What he said:

‘Through selection of immune escape viruses in the laboratory under appropriate containment conditions, we were able to identify the key regions [that] would enable 2009 H1N1 viruses to escape immunity,’

Now recall that the H1N1pdm 2009 virus is NOT a particularly nasty variant; that it has NOT been proved the escape mutants will infect vaccinated people at all – and that all the work was done "a state-of-the-art laboratory at the Institute for Influenza Virus Research in Madison", so the odds that it will get out are VERY low.

But papers have been sold, and the scare is in.

Internet archaeology

28 June, 2014

There I was, innocently doing a search on

Search the BIONET methods/bionet.molbio.methds-reagnts

- because my own “Manual of Online Molecular Biology Methods” links to it, and I was a regular there when it was operational – when I typed my own name in (as one does), and discovered that I had written this, as an answer to a post of 21 years ago.  Can’t even remember doing it.

 

S Poidinger writes:
> Gimme a K...Gimme an I....Gimme a T....whatdoesitspell?

- and I am prompted to reply:

Well, come on all of you bright young men
ProBoehAmPharm needs your help again
Got itself in a bit of a jam -
Stock price falling in the US of Am
So put down your books, pick up a kit
Gonna do a whole lot of sh*t...

And it's 1, 2, 3
What are we working for
Don't ask me, I don't give a sh*t
Long's I can use a kit

And it's 5, 6, 7
Open up the Plastic box
Ain't no cause to wonder why
ProBoehAmPharm ain't gonna die....

- With apologies to Country Joe and the Fish

A new virus from the Namib – and a guilty secret revealed

26 June, 2014

I have to confess to a guilty secret: there is a pleasure-inducing activity I have been indulging in for a week at a time these past three years.

And not alone….

This consists of going to the Gobabeb Research & Training Centre in the Namib Desert as part of an international “scientific expedition” aimed at investigating microbial soil biodiversity in the sandy and stony desert round Gobabeb.  These were started by Professor Don Cowan when he was at the University of the Western Cape, and have fortunately continued now that he has moved to a new Institute at the University of Pretoria.

Typical quartz-associated hypolith

Typical quartz-associated hypolith

I put the scientific expedition in quotes because anything that much fun shouldn’t be called scientific, but hey, it’s already resulted in one major paper on hypolith-associated viruses that I’m a minor author on, another co-authored opinion piece in the South African Journal of Science on biodiversity assessment that got the cover, as well as me being invited to be part of the Gobabeb station’s Microbiology & Fungi research “Theme Group“.

Moreover, I am now on the Board of the Institute for Microbial Biotechnology and Metagenomics at UWC on the strength of working with Marla Trindade and Lonnie van Zyl and others on scraping bits of green stuff off rocks and then watching them ultrafilter washings of it – so I suppose that we really did do some science, even if it was sinfully enjoyable. In any case, something that happened last year took me back to my roots – as well as possibly getting me some street (or gully) cred with the biodiversity crowd.

The heavily gullied area near Homeb

The heavily gullied area near Homeb

Basically, there we were in the Welwitschia-rich gullies near Homeb, 11 km from Gobabeb, visiting said plants.  There had been some rain 3 weeks previously, apparently, and there was an amazing eruption of foliage from some kind of bulb, every plant the same age and every one frantically flowering for all they were worth.  This alone was noteworthy, as the gullies were completely devoid of any trace of such plants the previous year. As we were wandering about, looking at Welwitschias, one Olivier Zablocki from the Univ Pretoria team – who had just done a MSc in plant virology with Gerhard Pietersen at UP – said something along the lines of “I wonder if there are any viruses infecting these plants?”.

welwitschia 2012

Welwitschia down in the gully

“What, like that one?” I said, having just fortuitously noticed a plant with tell-tale streaks on its leaves.  Of course, I seem to have lost my photos – temporarily, I hope! – after a Mac Mini OSX update disaster, but Olivier was kind enough to provide the necessary:

diseased albuca

Streaky Albuca. Note how quickly the fruit has formed, just a couple of days after flowering.

This sparked a flurry of activity, with people being called to observe the plant, and going out and looking for more.

Which were not found: not one other plant, of the hundreds we saw there and nearer Homeb, had any streaks at all.  What is more, they were growing all up and down some of the more inhospitable gravelly and rocky slopes I have ever seen, meaning they had to be seriously drought-tolerant, given the unlikelihood of them ever being exposed to much water.  This meant they must be ephemeral, or putting out foliage and flowers only after rain – and I have never seen anything flower as fast; three days later they were already fruiting.

albuca 2013

Albuca growing in the gully

We made the collective decision that this was sufficiently scientifically interesting to warrant its collection, and the plant was carefully dug up – with difficulty; the onion-like bulb was deep and seriously embedded among rocks – and carefully transported back to Gobabeb, hopefully for identification and then packaging to be taken back to Pretoria.

healthy albuca

Healthy Albuca growing in gravel

And yes, we did have a permit!

Meaning the foliage could go back to Pretoria, and there be subjected by Olivier to electron microscopy, and then RNA isolation and cDNA synthesis.  And lo, it came to pass – that a new potyvirus was discovered.  Kudos, Olivier and the Cowan lab!  The ms is submitted, and we wait only for…well, acceptance would be nice, but the proof is in the sequence.  And the pictures – murky EMs done by Olivier from precious tissue extracts, to boot.

Transmission EM from diseased Albuca extract

Transmission EM from diseased Albuca extract

And it took an old plant virologist to find it.  Life in the greying dog yet!

…and now it’s published – as a Disease Note in Plant Disease.  Thanks Evelien!

HPV type 16 E7 protein bodies cause tumour regression in mice

26 May, 2014

See on Scoop.itIIDMM News

Background

Human papillomaviruses (HPV) are the causative agents of cervical cancer in women, which results in over 250 000 deaths per year. Presently there are two prophylactic vaccines on the market, protecting against the two most common high-risk HPV types 16 and 18. These vaccines remain very expensive and are not generally affordable in developing countries where they are needed most. Additionally, there remains a need to treat women that are already infected with HPV, and who have high-grade lesions or cervical cancer.

Methods

In this paper, we characterize the immunogenicity of a therapeutic vaccine that targets the E7 protein of the most prevalent high-risk HPV – type 16 – the gene which has previously been shown to be effective in DNA vaccine trials in mice. The synthetic shuffled HPV-16 E7 (16E7SH) has lost its transforming properties but retains all naturally-occurring CTL epitopes. This was genetically fused to Zera(R), a self-assembly domain of the maize gamma-zein able to induce the accumulation of recombinant proteins into protein bodies (PBs), within the endoplasmic reticulum in a number of expression systems.

Results

High-level expression of the HPV 16E7SH protein fused to Zera(R) in plants was achieved, and the protein bodies could be easily and cost-effectively purified. Immune responses comparable to the 16E7SH DNA vaccine were demonstrated in the murine model, with the protein vaccine successfully inducing a specific humoral as well as cell mediated immune response, and mediating tumour regression.

Conclusions

The fusion of 16E7SH to the Zera(R) peptide was found to enhance the immune responses, presumably by means of a more efficient antigen presentation via the protein bodies. Interestingly, simply mixing the free PBs and 16E7SH also enhanced immune responses, indicating an adjuvant activity for the Zera(R) PBs.

 

I thank Russell Kightley Media for use of the HPV/cervical cancer graphic

Ed Rybicki‘s insight:

I keep saying – you gotta go green…B-) And here we are, suiting action to words.  

Modestly, of course.  

Well done to Mark Whitehead and Thomas Oelschlager; thanks to Inga for sticking with a difficult ms – and thanks Era Biotech for the technology!

See on www.biomedcentral.com

Virus experiments risk unleashing global pandemic, study warns

21 May, 2014

See on Scoop.itVirology News

Benefits of scientific testing in the area are outweighed by risks of pathogenic strains spreading round world, say researchers

Ed Rybicki‘s insight:

…and others say "Rubbish!"  I particularly like this bit:

 

"[Ron] Fouchier said…the authors had misinterpreted published data to arrive at their risk of someone picking up a virus in the laboratory. "The truth is that scientific research has never triggered a virus pandemic.""

The report goes on to say:

"Lipsitch and Galvani point out that a flu strain that spread around the world from 1977 to 2009 was probably released in a laboratory accident."

Yes.  But.  That was from the old Soviet Union – and they had a number of nasty things escape from laboratories, including anthrax and weaponised smallpox, by all accounts.

 

See on www.theguardian.com


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