Posts Tagged ‘Congo’

The Discovery of Filoviruses

10 March, 2015

Ebola is 40 today!

The discovery of filoviruses: Marburg and Ebola

Marburg virus

In 1967, the world was introduced to a new virusthirty-one people in Marburg and Frankfurt in Germany, and Belgrade in the then Yugoslavia, became infected in a linked outbreak with a novel haemorrhagic fever agent. Twenty-five of them were laboratory workers associated with research centres, and were directly infected via contact with infected vervet monkeys (Chlorocebus aethiops) imported to all three centres from Uganda.  Seven people died.   In what what was a remarkably short period of time for that era – given that this was pre-sequencing and cloning of nucleic acids, let alone viruses – it took less than three months for scientists from Marburg and Hamburg to isolate and characterise what was being called “green monkey virus” virus. The new agent was named Marburg virus (MARV), after the city with the greatest number of cases.

The first electron micrograph of the virus clearly exhibits the filamentous nature of the particles, complete with the now-famous “shepherd’s crook”.

The virus disappeared until 1975, when an Australian hitchhiker who had travelled through what is now Zimbabwe was hospitalised in Johannesburg, South Africa, with symptoms reminiscent of Marburg disease. He died, and his female companion and then a nurse also became infected with what was suspected to be yellow fever or Lassa viruses. In an example for later outbreaks, this led to rapid implementation of strict barrier nursing and isolation of the patients and their contacts, which resulted in quick containment of the outbreak – with recovery of the two secondary cases. MARV was later identified in all three patients.

Ebola virus

Ebola viruses burst from obscurity in 1976, with two spectacular outbreaks of severe haemorrhagic fever in people – both in Africa. In the better-known outbreak for which the viruses were later named, Ebola virus (EBOV) was first associated with an outbreak that eventually totalled 318 cases, starting in September 1976.  This was in the Bumba Zone of the Equateur Region in the north of what was then Zaire, and is now the Democratic Republic of the Congo (DRC).  The index case in the outbreak, as well as many of those subsequently infected, was treated in the Yambuku Mission Hospital. He was injected with chloroquine to treat his presumptive malaria: within a few days fever symptoms developed again; within a week, several others who had received injections around the same time also developed fevers which in several cases had haemorrhagic complications. 

Interestingly, women 15-29 years of age were most affected by the disease: this was strongly correlated with their attending antenatal clinics at the hospital, where they regularly received injections.

Apparently the hospital had only five old-style syringes and needles, and these were reused without proper sterilisation.  Nearly all cases in this outbreak either received injections at the hospital, or had close contact with those who had. 

Most people were infected within the first four weeks of the outbreak, after which the hospital was closed because 11 of 17 staff had died.  Another  269 people died, for a total estimated case-fatality rate of 88%.

The incubation period for needle- transmitted Ebola virus was 5 to 7 days and that for person to person transmitted disease was 6 to 12 days.

Interestingly, in post-epidemic serosurveys in DRC, antibody prevalence to the “Zaire Ebola virus” has been 3 to 7%: this indicates that subclinical infections with the disease agent may well be reasonably common.

The team that discovered the virus at the Antwerp Institute of Tropical Medicine in Belgium, did so after receiving blood samples in September 1976 from a sick Belgian nun with haemorrhagic symptoms who had been evacuated from Yambuku to Kinshasa in the DRC, for them to investigate a possible diagnosis of yellow fever.  Following her death, liver biopsy samples were also shipped to Antwerp – where the team had already ruled out yellow fever and Lassa fever.  Because of the severe nature of the disease, and its apparently novel agent, the World Health Organisation (WHO) arranged that samples be sent to other reference centres for haemorrhagic viruses, including the Centres for Disease Control (CDC) in Atlanta, USA.

The Belgian team were the first to image the virus derived from cell cultures on an electron microscope – when it was obvious that the only thing it resembled was Marburg virus. 

Image copyright CDC / Frederick A Murphy, 1976

Image copyright CDC / Frederick A Murphy, 1976

The CDC quickly confirmed that it was Marburg-like, with possibly the most famous virus image in the world, but that it was a distinct and new virus.

This meant it needed a name – and it was given one derived from the Ebola River that was supposed to be near the town of Yambuku.

Google map of the area where the first Ebola haemorrhagic fever outbreaks occurred

Google map of the area where the first Ebola haemorrhagic fever outbreaks occurred

Another, minor outbreak of the virus occurred in June 1997 in Tandala in north-western DRC: one young child died, and virus was recovered from her – and subsequent investigations showed that “two previous clinical infections with Ebola virus had occurred in 1972 and that about 7% of the residents had immunofluorescent antibodies to the virus”. This further reinforced the idea that subclinical infections were possible.

Sudan virus

In June 1976 – before the Yambuku epidemic in DRC –  an outbreak of a haemorrhagic fever began in the southern Sudanese town of Nzara.  The presumed index case was a storekeeper in a cotton factory, who was hospitalised on June 30th, and died within a week.

There were a total of 284 cases in this outbreak: there were 67 in Nzara, where it is presumed to have originated, and where infection spread from factory workers to their familes.  There were also 213 in Maridi, a few hours drive away – where, as in Yambuku, the outbreak was amplified by “nosocomial” or hospital-acquired transmission in a large hospital. In this case, transmission seems to have been associated with nursing of patients.  The incubation period in this outbreak was 7 – 14 days, with a case mortality rate of 53%.

Two viral isolates were made from sera from Maridi hospital patients in November 1976. Antibodies to the now-identified “Ebola virus” from DRC were detected in 42 of 48 patients clinically-diagnosed patients from Maridi – but in only 6 of 31 patients from Nzara.  However, it was subsequently shown that the Sudan and DRC Ebola viruses were different enough from one another to be separate viral species (see later), which undoubtedly affected the results.

Interestingly, 19% of the Maridi case contacts had antibodies to the virus – with very few of them with any history of illness.  This strongly indicates that the Sudan virus can cause mild or even subclinical infections.

An indication of the possible origin of the epidemic is the fact that 37% of the workers in the Nzara cotton factory appeared to have been infected, with 6 independently-acquired infections – and that this was concentrated in the cloth room, where there were numerous rats as well as thousands of insectivorous bats in the roof.  However, subsequent study of antibodies in the bats failed to detect evidence of infection, and no virus was isolated from bat tissue.

There was another outbreak of the same type of Ebola haemorrhagic fever in the area of Nzara in July – October 1979: this resulted in 34 cases, 22 of them fatal, with the index patient working at the cotton factory and all others being infected via the hospital he was admitted to.  It is interesting that antibodies to the Sudan virus were detected in 18% of adults not associated with the outbreak, leading the report’s authors to speculate that the virus was endemic in this region.

It was thought that the Sudan and DRC outbreaks were linked: the original WHO Bulletin report on the Sudan outbreak even speculates that extensive truck-borne commercial goods traffic between Bumba in DRC and Nzara in what is now South Sudan could have caused the DRC outbreak.  However, comparisons between the viruses isolated from the two epidemics later showed that they were distinct, both in terms of virulence, and antigenicity – meaning the Sudan virus got its own name.

Epidemics and outbreaks have resulted from person to person transmission, nosocomial or in-hospital spread, or laboratory infections. The mode of primary infection and the natural ecology of these viruses are unknown. Association with bats has been implicated directly in at least 2 episodes when individuals entered the same bat-filled cave in Eastern Kenya. Ebola infections in Sudan in 1976 and 1979 occurred in workers of a cotton factory containing thousands of bats in the roof. However, in all early instances, study of antibody in bats failed to detect evidence of infection, and no virus was isolated form bat tissue.

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Monkeypox vaccine?? We don’t need no monkeypox vaccine….

22 December, 2011

An in-press article in Vaccine that was tweeted by MicrobeTweets (well worth signing up to, BTW) has the intriguing title “Whither monkeypox vaccination?”

Now, some background to this: monkeypox virus is a rather nasty relative of smallpox (family Poxviridae; subfamily Chordopoxvirinae, genus Orthopoxvirus), meaning it is a large dsDNA virus (170-250 kb) with a complex structure.  The virus is endemic in remote forest areas in central Africa – principally in the Democratic Republic of the Congo – and naturally infects a number of animal species, including giant pouched rats (Cricetomys sp.), dormice (Graphiurus sp.) and African squirrels (Heliosciurus, Funisciurus), as well as laboratory monkeys, which is how it was isolated and got its name.

Monkeypox gets transmitted to humans by contact with infected animals: this includes by simple handling, as well as by exposure to meat and blood of butchered animals.  It causes a disease in humans that is very similar in appearance to smallpox, with a case fatality rate of 1-10%, but is apparently far less easily transmitted person-to-person.  It caused only sporadic and limited outbreaks in Africa and was of limited interest until an outbreak in the USA in 2003, which was linked to young prairie dogs kept in a pet store in close proximity to an infected Gambian pouched rat (Cricetomys gambianus) recently imported from West Africa. Seventy-three people were reportedly infected, among whom there were no fatalities.  The CDC recommends vaccination of people exposed to human or suspected animal cases with smallpox vaccine, as this protects animals from experimental lethal monkeypox challenge.

The Vaccine paper makes the point that the potential for monkeypox virus (MPX) to fill the disease niche recently vacated by smallpox was evaluated in the 1970s – and discounted, largely because human-to-human spread was inefficient enough for outbreaks not be self-sustaining – thus, although smallpox vaccine protected against MPX, the WHO thought there was insufficient justification to continue vaccination.

Now, however, the incidence of the virus in humans

“…appears to have markedly increased. In addition to diminished vaccine-induced orthopoxvirus immunity, there have been profound social and demographic changes that have increased human MPX exposures and the likelihood of severe disease. Recurrent civil war and subsequent economic decline have forced rural residents to flee deep into the rain forests for extended periods of time, disrupted traditional village life and increased dependence on hunting for sustenance, thus increasing exposure to animal reservoirs of MPX.”

So, in other words, people are getting a whole lot more exposure to sick animals.  Increasingly, by eating them.  The paper goes on to say:

“Although orthopoxviruses are relatively genetically stable MPX has diverged into two clades with different levels of virulence. As incidence rises, each new MPX infection provides an opportunity for viral evolution or adaptation that may result in a more virulent or contagious variant capable of sustained person-to-person transmission. These new circumstances merit a re-evaluation of the need for immunizing against MPX”.

So – that should be relatively simple, surely?  I mean, South Africa alone has millions of doses of smallpox vaccine safely frozen away from the 1970s?  Not so fast….

“However, in an era where the threat of smallpox is not imminent and there are conditions such as AIDS, tissue transplantation, and therapies for cancer and autoimmunity that cause immunodeficiency, the adverse events associated with live vaccinia are no longer considered acceptable for the general population.”

The paper goes on to mention how all sorts of supposedly safe new smallpox vaccines have been deposited into biodefence stockpiles, based on animal testing.

And there it is again – that word “biodefence”, in the context of human vaccines – implying that there is a “biothreat” to counter.  Specifically, in this case, the spectre of weaponised smallpox.

The authors go on to make reasonable statements about surveilling for monkeypox in central Africa, and vaccinating people at risk, and say that treatment options should also be investigated given that clinical diagnosis is relatively easy.

They also close with this:

“If immunization studies in developing countries are contemplated to support the licensure of orthopoxvirus vaccines for industrialized countries or for military purposes, then provisions from those countries or organizations should be secured to distribute successful products in endemic regions where the products were tested.” [my emphases]

I should hope so.  I should really, really hope so – because then one country’s biodefence interests could end up benefitting quite a few others, who are the ones who really need the product.  Now, while you’re busy with that, what about vaccines for Rift Valley fever, Crimean-Congo haemorrhagic fever and Chikungunya – which are actually far more serious a problem, in a much bigger geographical area?


HIV: roots run deeper than we knew

2 October, 2008

I have previously posted a number of articles on “molecular archaeology” of viruses, and how one can use extant sequences, archived tissue samples, or even blood of pandemic survivors to speculate on the origins of specific viruses, of viruses generally, or on the nature of old pandemic strains.Now HIV falls under the spotlight – again – as the 2nd October issue of Nature publishes three articles (one letter, a commentary on it and an independent commentary) on the origins of HIV-1 pandemic strains.I picked up on the first news – evidence for an older-than-previously-thought origin for HIV-1 – via our local paper this morning. Now this is VERY impressive; they usually keep science news for a slow day, and here they were telling us about a Nature paper on the day it was published! Accessing Nature brought up the Nature News commentary by Heidi Ledford, titled “Tissue sample suggests HIV has been infecting humans for a century”. Essentially, the commentary summarises the findings of Michael Worobey of the University of Arizona and his colleagues, who managed to amplify and sequence HIV-specific cDNA and DNA from a paraffin-embedded lymph node biopsy dating from 1960 from a woman in Léopoldville (now Kinshasa) in what is now the DR Congo. To quote Ledford:

“Their results showed that the most likely date for HIV’s emergence was about 1908, when Léopoldville was emerging as a centre for trade.”

Their findings added credibility to an earlier demonstration of HIV-1 in a 1959 sample, also from Kinshasa. What was interesting was that the sequences of the two viruses differed by 12%: this indicates that there was already significant divergence in the HIV-1 strains infecting people as early as 1960, pointing to a longer history of human infection than the previous estimate of the 1930s.Which led on to the Comments section, where one finds gems like this:

“This is one of the most stupid discovery I have ever heard. You will blame every single human plague on Africa, This is against all the Theories of evolutionary biology where The descents of the people that lived in the area might have developed a kind of resistance instead of being vulnerable to a new strain of the Virus.”


“HIV is older than your great-grandparents, uh-huh! And I’ll bet that the US bio weapons effort is just ecstatic about this deflection. So now these members of science play to the bio-jackboot population controllers with this ‘revelation’ that those sex-crazed Africans of course just couldn’t stop themselves from pulling chimpanzees (I thought the original scientific theory was “green monkeys”) out of the trees for a quickie.”

I couldn’t take this, so I replied:

“It continues to amaze me, as a teacher of virology who tells big classes every year where HIV comes from, how every year some clique of students takes the African origin of HIV personally, as a direct affront. I echo the correspondent above: it is a virus, people. Viruses infect animals, they infect people, and sometimes spread from one to the other – and back, if you are a zoo animal and catch something from your handler. The AIDS pandemic is an accident of sociology, demography, access to high-speed, long-distance travel – and truck routes, and truck drivers. It happens that it originated in Africa. So did the human race – only a lot longer ago. Inevitably, as humans encroached on apes, things get passed across. And don’t spread, much, until…someone puts a road through the village.Why don’t people get more exercised about the origins of HTLV, another retrovirus that almost certainly jumped from monkeys to humans? Except that happened many thousands of years ago, and in south-east Asia, not Africa. And for the same reasons: people eat monkeys and great apes. For that matter, it is speculated that chimpanzees got SIV-CPZ from vervet (I HATE the term “green”) monkeys – and that it may have caused a population bottleneck, some 100 000 years ago. I note that chimpanzees are known to eat vervets, incidentally – so they caught the virus the same way we did.

Ah, well…. In any case, Paul Sharp of the University of Edinburgh – and phylogenetics guru – and the godmother of HIV/SIV diversity, Beatrice Hahn of the University of Alabama (from whom I got the chimp-vervet virus link), have an independent commentary in the same issue, wherein they speculate on “The prehistory of HIV-1”. They make this very interesting comment:

“If the epidemic grew roughly exponentially from only one or a few infected individuals around 1910 to the more than 55 million estimated to have been infected by 2007, there were probably only a few thousand HIV-infected individuals by 1960, all in central Africa. Given the diverse array of symptoms characteristic of AIDS, and the often-long asymptomatic period following infection, it is easy to imagine how the nascent epidemic went unrecognized.”

They also make the important point that the findings of the Worobey group were replicated – with similar but non-identical virus sequences being found – by another group working independently with the same tissue sample. This is important because it nails down the findings more firmly, as HIV sequences within an individual do differ, and:

“…the distance along the evolutionary tree from the group M ancestor to the ZR59 or DRC60 sequences is much shorter than those between the ancestor and modern strains, consistent with the earlier dates of isolation of ZR59 and DRC60, and confirming that these viruses are indeed old”.

a, The HIV-1 genome fragments that were successfully amplified from DRC60 (red) and are available for ZR59 (black). The numbering for the HIV-1 sequences corresponds to the HXB2 reference sequence (Supplementary Table 1). b, The A/A1 subtree from the unconstrained (in which a molecular clock is not enforced) BMCMC phylogenetic analysis. 1960.DRC60A is the University of Arizona consensus sequence, and 1960.DRC60N is the Northwestern University consensus sequence (that is, the sequences independently recovered in each of the two laboratories). The DRC60 sequences form a strongly supported clade with three modern sequences also sampled in the DRC.

Reproduced with permission from Nature Publishing Group (RightsLink License No 2041420001096) from:
Direct evidence of extensive diversity of HIV-1 in Kinshasa by 1960
Michael Worobey, Marlea Gemmel, Dirk E. Teuwen, Tamara Haselkorn, Kevin Kunstman, Michael Bunce, Jean-Jacques Muyembe, Jean-Marie M. Kabongo, Raphaël M. Kalengayi, Eric Van Marck, M. Thomas P. Gilbert & Steven M. Wolinsky
Nature 455, 661-664(2 October 2008) doi:10.1038/nature07390

So where did the virus infecting humans come from? The best guess, from the paper and the commentaries, is that it originated – as do the extant chimpanzee virus supposed to have descended from the common origin – in chimpanzees somewhere in southeast Cameroon.How did it get into people? Sharp and Hahn again:

“The simplest explanation for how SIV jumped to humans would be through exposure of humans to the blood of chimpanzees butchered locally for bushmeat.”

No sex, no weird practices…just eating our cousins.  And how and why did it get to Léopoldville? Trade…and in those days before widespread truck routes, that would have been via rivers – which, Sharp & Hahn point out, drain from southeast Cameroon into the Congo River, which flows past what is now Kinshasa. The Worobey paper has some interesting history in it, documenting times of founding and rates of growth of cities in equatorial west Africa: Léopoldville/Kinshasa was and probably still is by far the fastest-growing of these, and was the earliest founded (in 1885). All that was needed to seed a pandemic, then, was that people infected by a virus as a result of butchering chimpanzees, moved some 700 km down natural trade routes to an emergent trade centre – and settled, and passed it on.Then, of course, it is the same old story, told so well by Jared Diamond in “Guns, Germs and Steel“: increased human population density and breakdown in social structure leads to increase in rate of transmission and incidence / prevalence of a disease agent, until it reaches the threshold necessary to break out. It is interesting that it took so long to become noticed – but then, HIV is passed on considerably less efficiently than Hepatitis B virus, so the pace of the epidemic was necessarily slow.But very sure….