P2 wild viruses, India

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P2 wild viruses, India

• EMRO Labdirectors Meeting
• Amman, 24-28 August 2003

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Issues

• Last P2 wild virus - Oct 1999, India
• ERC Mumbai detected 9 P2 wilds in late 2002/early
  2003, 7 x AFP, 1x contact, 1 x environment
• Lab strain, MEF-1, identified in 9/9 viruses
  sequenced
• Question - laboratory contamination or virus
  circulation in the population?

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P2 wild poliovirus (MEF 1)
12th Nov 02 6th Dec 02 Contact Sambhal, MRD
18th Dec 02 Kundarki, MRD
10th Nov 02 Mat, MTR
UP
16th Nov 02 1st Jan 03 Jawan, ALG
Gujarat
21st Jan 03 Kalyanpur, Jamnagar
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MEF-1

• A prototype wild P2, isolated in 1942 from
  soldiers in the Middle East Forces
• Originates from CNS from more than one fatal case
  (first inoculation in monkeys)
• Used as seed virus for IPV production
• Used as a lab strain in research studies
• Found once before in India - two samples in 2000
  from the Lucknow National Lab - lab contamination
  given as explanation

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Potential sources of lab contamination

• Use of reference MEF1 for research or as positive
  control in laboratory tests
• Recent experiments with MEF1 isolates from
  Lucknow 2000
• Anti polio 2-SL serum distributed by RIVM to ITD
  labs for ELISA found to be contaminated by live
  MEF1

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Investigations performed

• Site visit to ERC, Mumbai - concludes that lab
  contamination is very unlikely
• Most recent virus isolated in a National Lab
  that has no MEF-1 contaminated serum or MEF-1
  virus samples
• Re-isolation from original samples and sequencing
  of isolates obtained at RIVM confirm all ERC
  results

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Genetic investigations performed

• Sequence data show
• In 2000 and in 2002 two genetically different
  viruses involved, both present in prototype
  stocks of MEF1
• All isolates contain only one virus, but all
  viruses are different from each other
• Mutations do not add up in time

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Implications

• MEF1 present,but no circulation in population
• Multiple introduction of MEF1 in population
  during three months period, during which NIDs
  have taken place
• All AFP cases with MEF1 isolation were recently
  vaccinated
• Two scenarios for re-introduction of virus
• Unrecognised break in laboratory containment from
  a laboratory in India
• Most feared scenario contamination of vaccine

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Field investigation

• Government of India and WHO have installed an
  group of experts (JJ, WD, RS), many other persons
  have contributed to investigation
• Detailed investigation of all cases
• Epidemiological investigation around all cases
• Detailed analysis of all laboratory results
  (contamination in lab or during transport ?)
• Detailed investigation on the quality of
  surveillance activities in the region (cases
  missed ?)
• Identification of possible sources of MEF1 in the
  region (laboratories, universities, vaccine
  producers)
• Identification of links between cases and sources
  of MEF1

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Outcome of investigations

• Confirmation of all laboratory findings
• Vaccination of all AFP cases with OPV during
  NIDs, however in some cases after onset of
  symptoms
• Break of containment identified in one research
  laboratory, including a possible link to cases,
  but viruses present did not contain MEF1 !!!
• Unused vaccines vials of batches used in NIDs
  recovered from field

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Analysis of vaccins recovered from field

• Development of specific and sensitive PCR for
  detection of MEF1 in the presence of excess of
  three Sabin viruses (RIVM, CDC,NIBSC)
• Development of biological tests to detect
  presence of MEF1 in the presence of excess of
  three Sabin viruses
• RIVM use of Polyclonal NSL and SL antisera from
  ITD ELISA
• NIBSC Use of SL-specific monoclonal antibodies

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Sequence of amplificate is MEF1
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MEF1- marker VP1 position 99
MEF1- marker VP1 position 223
MEF1- marker VP1 position 300
MEF1- marker VP3 position 162
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Conclusions

• Vaccin lot 02586 from the field contains at least
  104 MEF1 particles
• MEF1 virus in OPV lot 02586 contains identical
  viruses to viruses present in stools from AFP
  cases
• Results within three days independently confirmed
  by NIBSC and later also by ERC, Mumbai (CDC
  primers)

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Next steps

• Intensive investigation of OPV vials from
  contaminated lot recovered from the field,
  producers and control agencies
• Testing of bulks of vaccines used to produce the
  contaminate OPV
• Test of lots and bulks produced just before and
  after incident and also of recent lots
• OPV from companies involved banned until end of
  investigation

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Next steps

• Intensive investigation ongoing
• Results are confidential