This document provides an overview of picornaviruses, with a focus on poliovirus. It begins by outlining the objectives of discussing picornavirus morphology, classification, pathogenesis, and the clinical manifestations, diagnosis, treatment and prevention of poliomyelitis and other diseases caused by echoviruses and rhinoviruses. It then provides details on picornavirus morphology, classification, the history of poliovirus research and discovery, epidemiology, characteristics, cultivation, pathogenesis, and clinical features of poliomyelitis. Key points covered include that poliovirus is an enterovirus that can cause paralysis, replication in the gastrointestinal tract and nervous system, and that infection may be asymptomatic, minor

1. PICORNAVIRIDAE
Dr.DineshKr Jain, MD., Assistantprofessor,
Department of Microbiology,SMS Medical
college, Jaipur

2. Objectives of today’s class
Toknow and understand the morphology,classification,
pathogenesis of Picornaviruses
Tolearn about the clinical manifestations, lab diagnosis,
treatment, prevention and prophylaxis ofPoliomyelitis.
Tolearn about the clinical manifestations, lab diagnosis,
treatment, prevention and prophylaxis of ECHOand
Rhinoviruses.

3. Introduction to Picornaviruses
Family Picornaviridae
Consists of a large number of very small RNAviruses,
27-30 nm insize
Resistant to lipid solvents like Ether, Chloroform, Bile salts

4. Morphology of Enterovirus

5. Morphology of Poliovirus
Electronmicroscopy(EM)ofPoliovirus

6. Epidemiology

7. Epidemiology

8. Classification
• 4 Genera pathogenic to humans:
1. Enterovirus – Infects intestinal tract
2. Rhinovirus – infects nasal mucosa
3. Hepatovirus
4. Paraechovirus
• 2 more Genera pathogenic for animals
1. Aphthovirus – Foot and Mouth disease of catlle
2. Cardiovirus – Encephalomyocarditis virus that infects mice

9. ENTEROVIRUSES- History
• Infantile paralysis – Paralytic disease of children
• Landsteiner and Popper (1909) demonstrated
Fatal case of Poliomyelitis –
Spinal cord &Feces inoculation –
Experimental disease transmission - monkeys

10. ENTEROVIRUSES– History Continued
• Enders, Wellers &Robbins (1949) demonstrated
• Growth of Poliovirus
• Culture of non-neural cells from human embryos
• Produced Cytopathic effects (CPE)
• Major break through – Nobel prize for this discovery
• Development of Virology (Milestone)

11. ENTEROVIRUSES– History Continued….
• Dalldorf &Sickles (1948) demonstration:
• Anew type of virus isolated
• From feces of children with paralytic poliomyelitis
• Named Type 1 Poliovirus
• This virus caused paralysis on inoculation in suckling mice
• Was called the Coxsackie virus
• Aspatient was from village ‘Coxsackie’in NewYork

12. ENTEROVIRUSES– History Continued ………
• Introduction of Tissue culture technique
• Diagnostic virology
• Led to Fecal isolation of several cytopathogenic viruses from cases
• Were called ‘Orphan viruses’
• Not associated with any particular clinical disease
• Was known by descriptive term “ECHO”
• ECHO– Entero cytopathogenic human orphan viruses

13. ENTEROVIRUSES– Characteristics
• Entero viruses are ‘Host specific’
• Infects only one or a few related species
• No common group antigen
• Antigen cross reactions observed in few

14. ENTEROVIRUSES– Classification
Group Serotype
Poliovirus (3) Type 1, Type 2, Type 3
Coxsackie virus A(24) 1-22, 24
Coxsackie virus (6) 1-6
Echovirus (34) 1-9, 11-27, 29-31
Numbered enterovirus
(EVsince 1969 – Numbered 68 andso)
68-71

15. Poliovirus
• Clinical case scenarion:
• A12 years old boy from Kovilpalayam presented to the emergency
department of KFMS&Rwith a h/o mild fever and sore throat X8
days, condition worsened and became severe accompanied by neck
rigidity and vomiting for last 2 days after a brief asymptomatic
period of 2 days.
• HOPI: On morning, boy experienced pain in lower limbs which
increased and progressed as weakness at the time of presentation.
• Mother’s history: Childhood (<5 years) vaccination not remembered

16. Poliovirus
• Diagnostic &Management strategy of Case scenarion:
• Throat swab, Stool, CSFsent to Diagnostic Microbiology Dept. for
viral studies
• Throat swab and stool specimen tested positive for Cytopathic effect
(CPE)in tissueculture.
• Virus was confirmed as Poliovirus type 1 by Neutralization test
• ReverseTranscriptase PCRwas positive for Poliovirus in CSF
• Patient improved on supportive treatment.

17. Poliovirus - Introduction
• Group IV;FamilyPicornaviridae; Genus Enterovirus; Species
Poliovirus
• The virus is composed of an RNAgenome and a protein capsule. The
genome is single-stranded positive-sense RNA genome that is about
7500 nucleotides long.
• Often called the simplest significant virus - First isolated in 1909by
Karl Landsteiner and Erwin Popper
• Egyptian paintings depicted the effects of polio by showing otherwise
healthy individuals with withered limbs.

18. Morphology
• Spherical Virion – Icosahedral symmetry
• 27 nm in diameter insize
• Composed of 60 subunits
• Consists of 4 Viral Proteins (VP1,VP2,VP3 &VP4)
• VP1 faces outside – Major antigenic site for combination
• VP1has type-specific neutralizing antibodies
• Viral Genome: Single stranded positive strand (ss RNA+ve sense)
• Virus can be crystallized – seen in cytoplasm of infected cells

19. Resistance
• Resistant to lipid solvents – ether, chloroform, bile, proteolytic
enzymes of intestinal contents and detergents
• Stable at a pH of 3
• In feces, it can survive for 4 months at 4ºC and for years at-20ºC
• Room temperature survival of virus in feces vary (one day to several
weeks) and It depends on temperature, moisture, pH and amount of
virus
• Readily inactivated by heat (55ºC X30 minutes)
• Molar MgCl2 , Milk or Icecream protects virus against heat
inactivation

20. Resistance
• Formaldehyde and Oxidising disinfectants destroy the virus
• Chlorination destroy the virus in water
• Organic matter present delays inactivation of virus
• Phenolic disinfectants not effective
• Does not survive lyophilisation well

21. Antigenic properties
• Based on Neutralisation test
• Poliovirus (PV)classified into 3 types: Type 1, Type 2 &Type 3
• Prototype strains:
Brunhilde &Mahoney strains for Type1
Lansing and MEFIfor Type2
Leon and Saukett for Type 3
Type 1 Most common, causes most epidemics
Type 2 Usually cause endemic disease
Type 3 Strains causeepidemic.

22. Antigenic properties
• Based on Complement Fixation Test(CFT)or Enzyme Linked
Immuno SorbentAssay (ELISA)or Precipitation tests
• Two antigens recognized: C&D
1. Cantigen - Capsid, Coreless, also called Heated or Hantigen
2. D antigen – Dense, also called Native or N antigen
• D antigen / N antigen associated with whole virion – Type specific
• Cantigen / H antigen associated with ‘empty’non-infectious virus
and Less specific and reacts with heterotypic sera

23. Antigenic properties
• D antigen – converted to Cantigen by heating the virus at 56⁰C
• Anti D antibody is protective. So,Potency of Injectable Polio Vaccine
can be measured in terms of D antigen units.
• Anti-C antibody does not neutralize virus infectivity

24. Host range &Cultivation
• Natural infection occurs only in humans
• Experimental transmission in monkeys by intracerebral or
intraspinal inoculation
• Chimpanzees and Cynomolgus monkeys – can be infected orally
• Established non-fresh strains can be grown in rodents, chick
embryos
• Virus grows readily in Tissue cultures of primate origin
• Primary Monkey Kidney cultures are used for Diagnostic purpose
and for Vaccination

25. Host range &Cultivation
• Cytopathic effect (CPE):Infected cells round up and become refractile
and pyknotic.
• Eosinophilic intranuclear inclusion bodies – may be demonstrated in
stained preparations.
• Well-formed plaques develop in infected monolayers with agar
overlay.

26. Pathogenesis
• The polio virus infects human cells by binding to an
immunoglobubin-like receptor called CD155 (poliovirus receptor).
• The exact mechanism that poliovirus uses for entering the cell is
unknown.
• However, the interaction of poliovirus and CD155 causes a change
in the shape of the viral particle that is needed to enter thecell
• There are two thesis' for the way the viral nucleic acid to enters the
cell.
1. RNAof poliovirus is injected into the host cell through a pore in the
membrane of the host cell.
2. Thepoliovirus is taken in by the host cell through endocytosis.

27. Pathogenesis
• Thegenome inside poliovirus can be used as mRNAand immediately
translated by the host cell.
• The poliovirus mRNAis then translated into a long polypeptide
which is cleaved into 10 individual viral proteins.
• Translation of the viral RNAoccurs by an IRES-mediated (internal
ribosome entry site) mechanism. The IRES is the extremely long 5’
end of the poliovirus’ mRNA. The assembly of viral particles is not
fully understood.
• The particles leave the host cell 4-6 hours after the initialinfection.
Each dying host cell can release 10,000 polio virions making
poliovirus lytic.

28. Pathogenicity
1. Virus transmitted by Fecal-oral route through ingestion. Other
possible modes in close contacts in patients of early stages:
Inhalation or Entry through conjunctiva of droplets of respiratory
secretions.
2. Virus multiplies initially in the epithelial cells of the alimentary
canal and in the lymphatic tissues, from the tonsils to peyer’s
patches
3. Spreads to regional lymph nodes and enters blood stream (Primary
viremia)
4. Further multiplication takes place in reticulo-endothelial system

29. Pathogenesis

30. Pathogenicity - Continues
5. Virus enters the Blood stream again (Secondary viremia)
6. Virus is now carried to Central Nervous System (CNS) Spinal cord
and Brain.
7. In CNS,Virus multiplies in selective neurons and destroys them
8. Earliest change: Degeneration of Nissl’s or bodies (Chromatolysis)
9. Nuclear changes follows.
10.When degeneration becomes irreversible, the necrotic cells lyses or
is phagocytosed by leucocytes or macrophages

31. Pathogenicity - Continued
11. Lesions are mostly in the anterior horns of the spinal cord causing
Flaccid paralysis, but posterior and intermediate horns can also be
involved.
12. Pathological changes will be more than distribution of paralysis
13. Encephalitis primarily involving the brain stem and extending up
to motor and pre-motor areas can occur insome cases.
14. Special circumstances: Direct neural transmission of Virus to
Central Nervous System (CNS) as in poliomyelitis following
Tonsillectomy through glossopharyngeal nerve present in
tonsillar fossa.
15. Poliomyelitis: Polio= gray matter; Myelitis= Spinal cord
inflammation

32. Clinical features
Poliomyelitis,orpolio,isacripplingdiseasecausedby
anyoneofthreerelatedviruses,poliovirustypes1,2or 3.
1. Inapparent infection
2. Minor illness
3. Paralytic poiliomyelitis or Major illness

33. Clinical features
Inapparent infection:
90-95% susceptible individuals develop only inapparent polioinfection
with Seroconversion alone.
Only 5-10% among them develop clinical infection. Incubationperiod:
About 10 days on average (Range:4 days –4weeks)
Minor illness:
Early manifestation is fever, headache, sore throat and malaise (Phaseof
primary viremia) lasting 1-5days called Minor illness or Abortive
poliomyelitis
Paralytic poliomyelitis or major illness:
Progression of infection 3-4 dyas after minor illness results in major
illness. Fever returns (Biphasic fever), along with headache, stiff neck and
other features of meningitis due to viral invasion of CNS(Polio case).

34. Pathogenesis

35. Poliomyelitis types and Complication
• Non- Paralytic polio: Disease does not progress beyond stage of
aseptic meningitis
• So,Types of polio: Nonparalytic polio &Paralytic polio.
• Paralytic polio can further be divided into: Spinal polio, bulbar polio,
and bulbospinal polio based on distribution of paralysis.
• Complication: Post-polio syndrome may also occur in which
symptoms ranging form breathing and swallowing problems to joint
pain, start many decades after the initial sickeness.
• Vaccine associated Poliomyelitis

37. Laboratory diagnosis
• Samples to be collected: Blood, CSF,Throat swab, Feces
• Transport: Immediately to lab in viral transport media (Hank’s
Balanced Salt Solution)
• Storage: 4⁰C (Days),-20⁰C(monthsto years)
• Laboratory Diagnostic tests available:
Viral isolation
Serodiagnosis
Molecular diagnosis

38. 1. Viral isolation
• In tissue culture – during primary Viremia 3-5 days afterinfection
before neutralizing antibodies appear (from blood)
• Early stages – isolation from throat swabs
• First week of infection: 80-85% viral isolation from feces
• Second week of infection: 50% viral isolation from feces
• Third week of infection: 25% viral isolation from feces
• Fecal excretion – intermittent (So two samples needed to be tested)
• Prolonged fecal excretion in immunocompromised, but no
permanent carriers,

39. 1. Viral isolation - continues
• Seldom poliovirus isolated from CSF,but it can be isolated from
spinal cord and brain, post-mortem diagnosis. (Unlike enteroviruses)
• Primary monkey kidney cells are employed commonly
• Human or Simian kidney cells can also be used.
• Inference: Viral growth indicated by Cytopathic effect (CPE)in 2-3
days.
• Mere isolation does not confirm the diagnosis

40. 2. Serodiagnosis
• Antibody rise appears after the onset of paralysis – demonstrated
by Neutralisation tests or Complement Fixation Tests (CFT).
• Neutralising antibodies appear early and persista for life
• In CFT,Anti-C antibodies appear first and disappear in few
months.
• Anti-D antibodies rise after few weeks and lasts for 5 years.
•CFTis useful to identify the exposure to Poliovirus but
not for type-specific diagnosis

41. 3. Molecular diagnosis
•Reverse Transcriptase PCR:
Viral RNAin CSFdemonstrated
•RNASequencing: In circulation 3 types of strains circulate
1. Wild virus
2. OPVstrain (Oral polio Virus)
3. VDPV– Vaccine Derived PolioVirus

42. Immunity
• Type-specificimmunity
• Humoralimmunity– circulatingandsecretoryantibodiesresponsible for
protectionagainstPoliomyelitis.
• IgM– appearin1weekandlastsfor6months
• IgG– appearby6monthsandpersistsfor life
• Neutralisingantibodiesprotectsagainstdisease
• SecretoryIgA– inGIT– providesIntestinalimmunity.

43. Immuno-prophylaxis

44. Immuno-prophylaxis
Liveoral polio vaccine (OPV) -four
doses in endemiccountries
or Inactivated polio vaccine (IPV)
given by injection - two-threedoses
depending on countryschedule

45. Immuno-prophylaxis

46. Immuno-prophylaxis

48. Immuno-prophylaxis
• UIP:BothIPVandOPVaresafe
• Salk(IPV)– Killedformolised,containstype1,2&3,S.cor Im
• Sabin(OPV)– LiveattenuatedvaccinegrowninMonkeykidneycells
• OPVhasType1virus10lakh,type2virus2lakh,Type3virus3lakh
• TCID50perdose(0.5mL)– vaccinemaintainedinair-tightcontainer
• PulsePolioImmunisation
• LocalIntestinalimmunity
• HerdImmunity

49. Treatment
• Currently there is no treatment to cure Polio. Treatment is focused on
supportive care. Moderate exercise - Anutritious diet
• Medication and rest to lower the fever and to reduce the painand
improve the strength. Breathing assistance with a ventilator
• ToPrevent Poliovirus: The most effective and most commonly used is
the Polio vaccine. This vaccine is given to young children in specific
increments.
• Vaccine works by strengthening and preparing the immune system
to a future encounter with the Poliovirus.

50. Coxsackie Group Avirus
• 24 serotypes
• Causes:
1. Herpangina (Vesicular pharyngitis),
2. Hand, Foot and Mouth disease (HFMD),
3. Aseptic meningitis,
4. Minor respiratory infection

51. Coxsackie Group Bvirus
• 6 serotypes
• Causes:
1. Epidemic pleurodynia or Bornholm disease
2. Myocarditis and pericarditis
3. Juvenile Diabetes – Coxsackie B4 ?
4. Orchitis
5. Transplacental and neonatal transmission
6. Post-viral fatigue syndrome

52. Coxsackie virus
• Labdiagnosis:
1. Animal inoculation – inoculating in suckling mice
2. Tissue culture – not useful
3. Serodiagnosis not practicable – due to several antigenic types

53. ECHOvirus
Enterocytopathogenic Human orphan virus
Grow in Human and simian kidney culture
Fever with rash, Aseptic Meningitis (most common cause)
Lab diagnosis: Feces, throat swab, CSF- Culture

54. New Entero virus
Acute hemorrhagic conjunctivitis – EV68,69,70,71
Radiculopathy – EV-70
Grows in Human embryonic kidney or HeLa cell lines.

55. Rhinoviruses
Common cold virus – 100 serotypes by neutralization
Transmitted by droplet infection
Culture – Just like for ECHO and other new entero viruses

56. Summary
• Polio virus – Belong to Entero virus – ss RNAvirus +sense
• Polio virus: 3 types – 1, 2 &3
• CD 155 receptor – Feco-oral transmission
• Paralytic or Non –paralytic polio
• CSF,Blood, Throat swab &feces
• Neutralisation tests, ELISA,CFT,RT-PCR
• OPV,IPV–Salk and Sabin
• Pulse polio immunization programme.

57. Takehome message
OPVandIPVtochuckoutPoliocompletely

58. Characteristics of Poliovirus
 PoliovirusisanEnterovirus. TrueorFalse?
 PoliovirusisaRNAvirus. TrueorFalse?
 Poliovirusistransmittedbydroplet nuclei. TrueorFalse?
 SalkvaccineisaKilled vaccine. TrueorFalse?
 SabinvaccineisaLiveattenuated vaccine. TrueorFalse?
 OralPolioVaccineisgivenassingledoseat birth. TrueorFalse?

59. References
• Ananthanarayan &Paniker’s Textbook of Microbiology, 9th Edition
• NAMH. How Polio Works. http://polio.emedtv.com/polio/polio-
treatment.html
• MayoClinic.com. Infectious Disease.
http://www.mayoclinic.com/health/polio/DS00572/DSECTION=sympt
oms
• Wikipedia. Poliovirus. http://en.wikipedia.org/wiki/Poliovirus
• Wikipedia. Poliomyelitis. http://en.wikipedia.org/wiki/Poliomyelitis

61. THANKYOU