Wednesday 6 January 2016

Zika virus, mosquitoes and a monkey on a platform...

As I noted yesterday the first published description of the isolation of Zika virus was from a study set up in the Zika forest in Uganda. It was described in print by Dr George Dick and colleagues in the September 1952 issue of Transactions of the Royal Society of Tropical Medicine and Hygiene.[1]

Rhesus macaque (Macaca mulatta) [2]
In the first study, which began April 1947, six platforms were set up in the forest canopy and upon each, a caged rhesus monkey was placed. 

The first Zika virus was identified from a monkey...

This was to be a Yellow fever virus (YFV) study since Zika forest was known to harbour a lot of previously YFV infected, antibody-positive monkeys-but stuff happens on the road to completing a good plan.

By the way: placing or penning an animal in a location known to, or under suspicion of, harbouring an infectious agent - a virus in this case - is the precess of creating a sentinel animal.

On 18th April, the daily temperature recording for "Rhesus 766" had increased to 39.7'C, rising to 40'C the next day. It was taken to the lab at Entebbe, a blood sample collected and then the primate was observed for the next month. The only sign or symptom of illness in 766 was the fever.

The blood was injected intraperitoneally (no signs of illness) or intracerebrally (became ill from day 10 post-injection) into mice and underneath the skin of Rhesus 771 (which did not develop any signs of illness).

A filterable "agent" (filters were used to remove everything but very small viruses) was recovered from the brains of the ill mice and also from the serum of 766 - it was later named Zika virus (766 strain). The virus's growth could be hampered by antibodies which developed in the serum of 766 a month later, identifying the that the Zika virus was capable of triggering a specific immune response, despite a mild illness. The more lab-savvy of you may have realised by now, that this all happened before cell culture was being used. The only way to "grow" or amplify more virus was to infect anew a susceptible host animal-which could also reveal whether the agent was capable of causing disease, so long as the experiments were suitably controlled.

A second Zika virus was found among mosquitoes...

A second virus (later called strain E/1) was acquired during a different study, but also set up for YFV. The E/1 was identified from a ground up, unfiltered preparation of Aedes africanus mosquitoes in saline which had been caught 11-12th January 1948 using a variation of the platform system. . The preparation was injected intracerebrally into 6 mice and subcutaneously into Rhesus 758 via 9 inoculations, across 2 weeks. 

Diagnosis of Zika virus by infection of animals...

Primate 758 showed no signs of illness, but the mice had - at the 7th day - so at day 8, 9 and 10 after 758's injection, blood was collected and serum injected intracerebrally into groups of 6 new mice. 
  • From the first primate sample (day 8), 2 injected mice died and virus could be passed on to additional mice from filtered preparations of their infected brain tissue
  • From the second sample (day 9), 1 mouse died (and its brain tissue filtrate could also produce new infections in mice). A second mouse was paralysed but another virus, called Theiler's mouse encephalitis virus [TMEV], a picornavirus, was identified upon further infections 
  • No mice died from the 3rd 758 inoculation of serum
  • The serum from 758 was shown to block infection of animals by Zika virus after it was first preincubated with preparations of:
    • the agent isolated from 758 serum after inoculation with the A. africanus preparation or
    • the E/1 Zika virus strain or
    • the 766 Zika virus strain
The authors concluded Zika virus was a new virus and that it triggered a specific antibody response which did not cross-react with YFV, dengue virus or the TMEV found in the paralysed mouse. 

It was also notable that disease was mild or went unnoticed in primates. Primates are usually considered to be close animal substitutes for humans in studies of disease progression. 

No studies of pregnant primates were conducted which does raise the question of whether we should consider a check-list of things to research for each and every virus capable of replicating in us.

  1. Zika virus. I. Isolations and serological specificity.
    Trans R Soc Trop Med Hyg. 1952 Sep;46(5):509-20.
  1. Added some links and worked out some typos and layout issues.

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