Enterovirus an underestimated infection

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Enterovirus -an underestimated infection

• Eva ampachová
• Virology - Hospital Ceské Budejovice
• 2nd Medical Faculty,
• Charles University, Prague

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Types of human enteroviruses
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Phylogenetic classification
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Clinical aspects of enterovirus infections
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Laboratory methods

• Culture
• various samples may be tested
• Cell cultures (monkey kidney, human embryonic
  cells)
• suckling mice (i.p., s.c, intracerebrally)
• RNA detection
• Serology
• neutralization test, labour intensive
• ELISA - heterotypic responses, persistence of IgM
• CFT - group reactive

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Methods used in our laboratory

• RT PCR -
• various samples, group specific primers
• protocol of Takami, based on Robards primers
• Cell culture
• monkey kidney, human embryonic fibroblasts
• Neutralisation test
• laboratory strains Cox. B1-6, patients strain
• Other serology
• rarely CFT, IFT with patients strain

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Results of investigation of CSFs
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Results of CSFs tested in 2003(total 42
patients)
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Results of CSFs tested in 2004 (total 148
patients)
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Results of CSFs tested in 2005(total 346
patients)
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Characteristic properties of the outbreak in 2003

• CSF findings
• high cell count (some gt1000 /3mm3 ), high
  proportion of neutrophils, protein increased,
  glucose normal
• Onset
• acute (similar to bacterial meningitis)
• Course
• short, up to 1 week hospital stay, relatively
  mild after the initial phase
• Long term sequelae
• none

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Charakteristic properties of the outbreak in 2004
- 5

• CSF findings
• Moderate to low cell count (30-750 /3mm3 ),
  predominantly lymphocytes, proteins increased,
  glucose normal
• Onset
• acute but not dramatic
• Course
• sometimes prolonged, some with transient paresis
• Sequellae
• rare, mild, none of them permanent

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Dual infections

• VIII 2004, M, 12 y., Enterovirus TBE
• XII 2004, F, 42 y., Enterovirus TBE
• XII 2004, F, 62 y., Enterovirus VZV
• IV 2005 , F, 6 y., Enterovirus EBV
• Cases of Enterovirus TBE infections had
  slightly more severe course than patients with
  single virus
• VZV Enterovirus infection has not differed from
  other Herpes zoster infections
• Enterovirus EBV infection was relatively mild
  meningitis

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Culture and typing

• 2003 - 3 strains
• Echo 33 and Echo 9
• 2004-5 winter - 12 strains
• 3x Echo 5 and Echo 30, 2x Echo 9 and Cox.B3, 1x
  Echo 7 and Cox. B1
• 2005 autumn - 5 strains
• 2x Echo 7, 1x Echo 5, Echo 30, Cox. B5

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Problems of strain typing

• Strains from one outbreak were defined as several
  different types
• Strain from close or family contacts were defined
  as different types
• Typing of enteroviruses is difficult and the
  Reference laboratory uses polyvalent sera
• We suggest serotyping of entrovirus is not always
  reliable

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Case 1

• Neonate, born in term (3200 g, 49 cm)
• Shortly after birth papulous exanthema emerged,
  later changed to vesicles.
• Baby was breast fed, had lower weight gain, but
  otherwise was in good condition
• HSV infection was expected, PCR for HSV 1, HSV 2,
  VZV was negative
• Localisation of exanthema was on hands and feet,
  small erosion in mouth was found

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Case 1

• Enterovirus was found from the vesicles and serum
• Exanthema and status improved in a week
• Baby was followed up but no sequellae were found

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Case 2

• In regular ultrasound screening of 32 years old
  woman in her 20th week of pregnancy, acquired
  heart damage was diagnosed in the foetus
• Destruction of the tricuspid valve, severe
  dilatation of the right ventricle, and
  haemodynamic disorder were found
• Results of genetic testing were normal
• Enterovirus was found in amniotic fluid by PCR
  and culture

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Case 2

• Enterovirus was identified as Echo 7
• Patient underwent termination of pregnancy
• In autopsy, foetal myocarditis was found,
  destruction of valves
• Enterovirus was detected in heart tissue of the
  foetus by PCR and culture
• Mother reported some respiratory infection
  approximately one month prior to the ultrasound
  screening

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Case 3

• In regular ultrasound screening of 24 years old
  woman in her 20th week of pregnancy, foetal
  hydrothorax and renal pelvis dilatation were
  found, yet no heart abnormality was found
• Patient underwent amniocentesis
• No genetic abnormalities of the foetus were found
• Enterovirus was proved by PCR and culture from
  amniotic fluid, type Coxsackie A 9

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Case 3

• Patient underwent termination of pregnancy
• In the autopsy of foetus pleural and pericardial
  inflammation with exsudate was found, no anatomic
  abnormalities were found
• Enterovirus was detected by PCR in pleural fluid
  
• No valid anamnestic data from mother were
  obtained (she was mentally impaired)

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Case 4

• Woman, 20 years old, with diagnosis of meningitis
  was admitted to hospital
• She was in her 17th week of pregnancy
• Tests for TBE were negative, enterovirus was
  detected in CSF by PCR
• Culture was not requested, strain for typing was
  not available
• Patient status improved in several days

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Case 4

• Amniocentesis was indicated
• No genetic abnormalities were found
• Enterovirus was not found (by PCR and culture)
• Risk of malformation of the foetus was set low
• Woman decided to continue with the pregnancy
• In term, she gave birth to a healthy infant

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Our spheres of interest and best co-workers

• Diagnosis of meningitis and other infectious
  diseases - collaboration with the Dept. of
  Infectious Diseases
• Collaboration with the Neonatology and Antenatal
  Clinic

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Collaboration not yet well established

• Collaboration with the Centre of Cardiology
• Collaboration with the Paediatric Clinic
• Collaboration with other specialists

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Conclusions I

• When number of testing increased, number of
  proved enterovirus infection increased
  proportionally, too.
• Enterovirus outbreaks occur from summer to autumn
  (may be extend to winter)

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Conclusions II

• Meningitis is the most often diagnosed
  enterovirus disease
• Mild infections will always be aside the interest
• Infection in pregnancy should not be
  underestimated
• More effort should be put in the investigation of
  inflammatory heart disease

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Conclusions III

• Not all clinicians are aware of the importance of
  the enterovirus infection
• Since the introduction of PCR, diagnosis became
  rapid and reliable
• Treatment is not yet available, some drugs are in
  clinical testing
• In principle, no broad-spectrum antivirals will
  be available

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Despite increasing number of investigations, real
number of enterovirus infections remains
unknown.Enteroviruses as a cause of serious
disease are often underestimated
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Acknowledgements

• N. Piskunova, M. Fialová, Laboratory of Genetic
  methods
• V. Chmelík and all his team, Dept. of Infectious
  Diseases
• M. Hanzl, J. Smrcka, Dept. of Neonatology
• H. tepanková, Advisor in Human Genetics
• D. Teislerová and all the team from our
  laboratory of virology