1
SPEAKER
J. B. Kathiriya
1040416004
Veterinary
Microbiology
Department of Veterinary Microbiology
College of Veterinary Science & A. H.,
Junagadh Agricultural University, Junagadh
MAJOR GUIDE
Dr. N. M. Shah
Rt. Professor & Head,
Dept. of Veterinary Microbiology,
Vet. College, SDAU, Dantiwada
MINOR GUIDE
Dr. J. S. patel
Professor & Head,
Dept. of Veterinary Medicine,
Vet. College, JAU, Junagadh

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Introduction and HistoryIntroduction and HistoryIntroduction and HistoryIntroduction and History
OvervieOvervie
ww
OvervieOvervie
ww
Animal Prion DiseaseAnimal Prion DiseaseAnimal Prion DiseaseAnimal Prion Disease
Function Normal PrionFunction Normal PrionFunction Normal PrionFunction Normal Prion
ConversionConversion ofof PrPc to PrPSCConversionConversion ofof PrPc to PrPSC
Pathogenesis of PrionPathogenesis of Prion
DiseaseDisease
Pathogenesis of PrionPathogenesis of Prion
DiseaseDisease
Human Prion DiseaseHuman Prion DiseaseHuman Prion DiseaseHuman Prion Disease
Prevention and ControlPrevention and ControlPrevention and ControlPrevention and Control
ConclusionsConclusionsConclusionsConclusions
Diagnosis and TreatmentDiagnosis and TreatmentDiagnosis and TreatmentDiagnosis and Treatment

INTRODUCTION
AND
HISTORY
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 PROTEINACEOUS + INFECTIOUS =PRION.
 Prion disease or transmissible spongiform encephalopathy
(TSE) are a group of neurodegenerative disorder.
 Smaller than smallest known virus.
 Devoid of nucleic acids.
 Not yet completely characterized.
 Manifested as infectious, genetic, and sporadic disorders.
 Accumulation of PrPsc the conformation of which differs
substantially from that of its precursor, PrPc.
 All prion diseases are fatal.
 There is no effective treatment.
 It is important to understand how these diseases are transmitted
in order to prevent their spread.
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Animal Prion DiseasesAnimal Prion Diseases
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7
(Bucelli et al., 2013)(Bucelli et al., 2013)
14%
85%
1%
Fatal insomnia
Human Prion Diseases

8
Tikvah Alper
Alper et. al., 1967
Exposed Brain of Scrapie infected mice to UV
radiation – could not alter the infectivity of
brain tissue.
History

9
Stanley B.Prusiner
The Noble Prize in Physiology or Medicine 1997 was awarded to
Stanley Prusiner “for his discovery of Prion-a new biological principal
of infection” in 1982.
www.nobelprize.org/prizes/medicine/1997/summary

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 1986 - Mad cow disease is first discovered in the United Kingdom.
From 1986 to 2001, a British outbreak affects about 180,000 cattle.
 January 1993 - The BSE epidemic in Britain reaches its peak with almost
1,000 new cases being reported per week.
 1996 - The first case of vCJD is reported. (Stephen Churchill -first victim)
 1996-1999 - The European Union bans British beef.
 1996 - Britain’s agriculture ministry confirms that mad cow disease can be
passed from cow to calf.
 2002-The first confirmed case of vCJD appears in the U.S., in a 22-year-
old British woman living in Florida.
 December 23, 2003 - The USDA confirms the first case of mad cow
disease in the United States.
 January 9, 2004 - The USDA destroyed about 130 cattle - first-ever US
case of mad cow disease.

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 September 5, 2008 - Canadian scientists announce a discovery
that paves the way for diagnostic testing of live cattle, rather than
postmortem.
 September 13, 2008 - An Alabama research study shows that mad
cow disease can sometimes be caused by genetic mutations.
 March 14, 2009 - The US government permanently bans the
slaughter of cows too sick or weak.
 March 2014 - After 15 years, the United States lifts the ban on beef
from the European Union, pending inspections.
 June 11, 2014 – The USDA announces recall of 4,000 pounds of
beef.
 March 24, 2016 - France confirms the first case of BSE since 2011.

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* Up to August, 2018
Number of cases of bovine spongiformNumber of cases of bovine spongiform
encephalopathy (BSE)reported in the UKencephalopathy (BSE)reported in the UK

GEOGRAPHIC DISTRIBUTION
Status of many countries is not known because they have no surveillance for
this disease
(CDC, 2018) 13
TMETME
TME has
also been
seen in
farmed
mink in
Canada,
Finland,
East
Germany
and the
former
USSR.

14
Indian scenario
Over the period spanning from 1968 to 1997, National Institute of Mental Health
and Neurosciences (NIMHANS), Bangalore, 69 cases of CJD recorded from
different parts of India.
New cases are emerging and many may have been unreported as no autopsies
were carried out.
From 1990 to 1998, Department of Neurology, G.B. Pant Hospital, New Delhi,
India admitted 10 cases of CJD from North India.
India has a population of more then one billion and assuming an annual
incident of 0.5 per million population, probably there would be at least
around 500 new cases of CJD annually in India.
Various studies have reported annual incidence of 0.5-1.5 cases
of CJD per million of general population.
(Mehndiratta et. al., 2016).

Normal protein
 Prion proteins normally present in CNS –called as
cellular proteins (PrPc
)
 PrPC
(C for cellular)
 The PrP protein is a sialo glycoprotein that occurs
on the surface of neurons and other cells.
 Primary structure dominated by alpha helix.
 Easily soluble
 Easily digested by proteases
 Encoded by PRNP gene (in humans)
 It’s function is not precisely known, But it may
play a role in cell adhesion, transmembrane
signaling or copper binding.
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16

Conversion of PrPc to PrPSC
The two mechanisms believed to give rise to PrPsc are:
1. Post-transcriptional modification leading to an
alteration in the conformation of the normal prion
protein
2. Mutations in the prion gene (PRNP) resulting in the
translation of a protein with sequence abnormalities
that predispose to the spontaneous adoption of an
abnormal conformation.
17

(Collinge and Clarke, 2007 ) 18

Abnormal Protein
 PrPSC (sc for scrapie)
 Same amino acid sequence and
primary structure as normal protein.
 Secondary structure dominated by
beta Plated sheet.
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Major difference between prion protein and its normal
counterpart is folding pattern of proteins.
Both isoforms encoded by the same cellular gene
PRNP, located on short arm of human chromosome 20.
Identical molecular weights of 33-35 kDa.
Amino acid sequences matches, but differ in their 3D
structure.
Major difference between prion protein and its normal
counterpart is folding pattern of proteins.
Both isoforms encoded by the same cellular gene
PRNP, located on short arm of human chromosome 20.
Identical molecular weights of 33-35 kDa.
Amino acid sequences matches, but differ in their 3D
structure.

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21

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23
Peripherally acquired
prions replicates in SLOs
Pathogenesis of Prion diseases

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Secondary lymphoid organs (SLOs), follicular dendritic cells (FDCs),
LTßR-Lymphoid tissue ß-receptor, TNF-Tumour necrosis factor,
Mfge8- Milk fat globule epidermal growth factor
Prion deposition occurs when PrPSC production exceeds to its clearance
Pathogenesis of Prion diseases
Autonomic nerves

(Jeffrey and González, 2007)
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Scrapie has been diagnosed in more than 1,000
flocks in the United States and has primarily
been reported in the Suffolk breed.
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27
27

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(Maddison et. al., 2010)

1. Scrapie in Pargana district of West Bengal
(Garole sheep) (2015)
2. Scrapie in Kamach sheep from Himalayan
region (1996)
3. Scrapie in Kasauli Himachal Pradesh (1996)
(Choudhary et. al., 2015)
29
Reports of Scrapie in Indian Sheep

2
3
8 8
10
8
6
4
2
0
KARNA
(J&K)
MANDHYA
Karnataka
MALPURA
Rajasthan
GAROLE
(WB)
(Chaudhry et.al., 2015) 30

Bovine Spongiform Encephalopathy (BSE)Bovine Spongiform Encephalopathy (BSE)
 First recognized among
cattle in the United
Kingdom in 1986.
 Cases may have occurred
in the early 1980s.
 Peaked in 1992-1993:
36,682 cases reported in
1992 alone.
 As of June 1st
2018,
1,84,400 total cases of
BSE in Great Britain.
 First recognized among
cattle in the United
Kingdom in 1986.
 Cases may have occurred
in the early 1980s.
 Peaked in 1992-1993:
36,682 cases reported in
1992 alone.
 As of June 1st
2018,
1,84,400 total cases of
BSE in Great Britain.
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BSE: U.K. Government Actions
33
Time course of epidemic BSE in the UK 1986-2000, with dates of major precautionary
interventions. Mammalian ban on meat and bone meal in March 1996 extended a 1994
ban for farmed food animal species to include all mammalian species. SBO = specified
bovine offals (brain, spinal cord, thymus, tonsil, spleen, and intestines from cattle >6
months of age); MBM = meat and bone meal (protein residue produced by rendering).
35

 BSE SURVEILLANCE
 A total of 350 suspected brain samples were
collected from Cross bred cattle (10),
buffaloes (315), sheep (21), and Goat (4).
 Not a single confirmed case is found.
(official IVRI Website)
36

(Novakofski et al., 2005) 37

 Consumption of beef and beef products.
 Lateral transmission is possible by taking bone meal
and blood meal of infected animals.
 Both vertical and horizontal transmission are
possible.
Placenta, fetal membrane, amniotic fluid may
contaminate the pasture and pen and surrounding
environment
(Pattison, et. al., 1972).
Dissemination of prion by ewes to their suckling
offspring through colostrum and milk
(Timm et. al., 2008). 38

SIGNS & SYMPTOMS
 BSE has a long incubation period : 2-8 years.
 Clinical signs of BSE are found in adult animals. Symptoms
may last for a period of 2-6 months before the animal dies.
 Animals with BSE may demonstrate some of the following
symptoms:
 nervous or aggressive behavior
 depression
 hypersensitive to sound and touch, twitching, tremors
 abnormal posture
 lack of co-ordination and difficulty in rising from a lying
position
 weight loss
 decreased milk production
37

Sampling
Obex, which is the portion of the brain that must be obtained for the
diagnosis of BSE and other spongiform encephalopathies such as
scrapie and chronic wasting disease.
(Center for Food Security and Public Health, Iowa State University, 2011)
38

Transmissible Mink Encephalopathy
(TME)
39
It causes neurological signs including
behavioral changes. The early clinical signs can
be difficult to analyzed, It may include
difficulty in eating and swallowing, and
changes in normal grooming behavior
(Center for Food security and Public health,
2016)

Chronic Wasting Disease (CWD)
 Known natural hosts:
Mule deer and White
tailed deer (Odocoileus
species) and Rocky
Mountain elk
 Clinical symptoms
 Weight loss
 Behavioral changes
 Excessive salivation
 Difficult swallowing
40
(Sigurdson and Miller, 2003)

Creutzfeldt-Jakob Disease (CJD)Creutzfeldt-Jakob Disease (CJD)
 Occurs worldwide
 According to the World Health Organization (WHO), over
220 cases of vCJD have been reported worldwide, with the
majority occurring in the UK (177 cases) and in France (27
cases).
 Because there is no way to detect BSE in blood, people who
have lived for long periods in areas where mad cow disease
has been found are not allowed to donate blood in the U.S.
 Annual incidence (U.S.): ~1 case per million population
 Median age at death (U.S.): 68 years
(Mead et al., 2003)
 Occurs worldwide
 According to the World Health Organization (WHO), over
220 cases of vCJD have been reported worldwide, with the
majority occurring in the UK (177 cases) and in France (27
cases).
 Because there is no way to detect BSE in blood, people who
have lived for long periods in areas where mad cow disease
has been found are not allowed to donate blood in the U.S.
 Annual incidence (U.S.): ~1 case per million population
 Median age at death (U.S.): 68 years
(Mead et al., 2003)
41

CJD – More Features
 Rapidly progressive - over 50% of CJD patients die
within 6 months and 85-90% within 12 months of illness
onset
 Median illness duration: 4 months
 Four different forms of CJD:
 Sporadic
 Familial
 Iatrogenic
 Variant form (v-CJD)-currently highly prevalent
 Rapidly progressive - over 50% of CJD patients die
within 6 months and 85-90% within 12 months of illness
onset
 Median illness duration: 4 months
 Four different forms of CJD:
 Sporadic
 Familial
 Iatrogenic
 Variant form (v-CJD)-currently highly prevalent
42

Sporadic CJDSporadic CJD
 About 85% of CJD patients
 No recognizable mode of transmission identified
 May be caused by:
 Age-related somatic mutation of the prion protein
gene
 Most cases of sporadic CJD occur in adults aged
between 45 and 75.
 On average, symptoms develop between the ages of
60 and 65.
 Despite being the most common type of CJD, sporadic CJD
is still very rare, affecting only 1 or 2 people in every
million each year in the UK.
 In 2014, there were 90 recorded deaths from sporadic CJD
in the UK.
 About 85% of CJD patients
 No recognizable mode of transmission identified
 May be caused by:
 Age-related somatic mutation of the prion protein
gene
 Most cases of sporadic CJD occur in adults aged
between 45 and 75.
 On average, symptoms develop between the ages of
60 and 65.
 Despite being the most common type of CJD, sporadic CJD
is still very rare, affecting only 1 or 2 people in every
million each year in the UK.
 In 2014, there were 90 recorded deaths from sporadic CJD
in the UK.
43

Familial CJDFamilial CJD
 About 10-15% of CJD patients
 Autosomal dominant inheritance
 Associated with prion protein gene abnormalities
 Usually a family history of CJD is present
 It affects about 1 in every 9 million people in the
UK.
 In 2014, there were 10 deaths from familial CJD and
similar inherited prion diseases in the UK.
 About 10-15% of CJD patients
 Autosomal dominant inheritance
 Associated with prion protein gene abnormalities
 Usually a family history of CJD is present
 It affects about 1 in every 9 million people in the
UK.
 In 2014, there were 10 deaths from familial CJD and
similar inherited prion diseases in the UK.
44

Iatrogenic CJD
 <1% of CJD patients
 Infection is accidentally spread from someone with CJD
through medical or surgical treatment.
 Transmission through:
 Human Growth Hormone
 Dura Mater Graft
 Cornea transplantation
 In 2014, there were 3 deaths from iatrogenic CJD in the UK
caused by receiving human growth hormone before 1985.
 <1% of CJD patients
 Infection is accidentally spread from someone with CJD
through medical or surgical treatment.
 Transmission through:
 Human Growth Hormone
 Dura Mater Graft
 Cornea transplantation
 In 2014, there were 3 deaths from iatrogenic CJD in the UK
caused by receiving human growth hormone before 1985.
45

-Variant CJD was first described in 1996 in UK.
 Has different clinical and pathologic characteristics from
other types of CJD.
 Till Feb 2006,159 cases of vCJD were diagnosed in UK
and 153 have died.
• Characterized by extreme rapid progression from symptoms
to disability and death.
Variant (vCJD)
Evidence for Relationship between BSE and vCJD
• Since 1996, evidence has been increasing for a causal
relationship between ongoing outbreaks of BSE and vCJD.
• Strong scientific evidences: agent responsible for BSE
(in cattle) and vCJD (in humans) is the same (PrPsc).
54

47

48
1. 1968 to 1997, National Institute of Mental Health
and Neurosciences (NIMHANS), Bangalore, 69
cases of CJD recorded from different parts of India.
2. vCJD in Karnatak and Kerala (1991)
vCJD in India
(Mehndiratta et. al., 2016)

Map presenting
Reported cases in
different states of India
Sporadic occurrence of
diseasesvCJD
vCJD
Scrapie
Scrapie
vCJD
57

50

Clinical and Pathologic Characteristics
Characteristic Classic CJD Variant CJD
Median age at death 68 years 28 years
Median duration of illness 4-5 months 13-14 months
Clinical signs and symptoms Dementia; early
neurologic signs
Prominent psychiatric/behavioral
symptoms; painful
dyesthesiasis; delayed
neurologic signs
Periodic sharp waves on EEG Often present Often absent
Pulvinar sign on MRI Not reported Present in >75% of cases
Presence of "florid plaques" on
neuropathology
Rare or absent Present in large numbers
Immunohitochemical analysis of
brain tissue
Variable
accumulation
Marked accumulation of
protease-resistance prion
protein
Presence of agent in lymphoid
tissue
Not readily
detected
Readily detected
Increased glycoform ratio on
immunoblot analysis of protease-
resistance prion protein
Not reported Marked accumulation of
protease-resistance prion
protein
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52
Death information was obtained from other surveillance mechanisms; data include familial
prion diseases. Rates are adjusted to the US standard 2000 projected population. 60
(CDC, 2018)

53
(Kirkwood et al., 1993) 61

54
Deaths from Kuru 1957-1982

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GERSTMANN-STRAUSSLER-SCHEINKER SYNDROME
(GSS)
• A familial disease with autosomal dominant inheritance,
• First described in1936 by Gerstmann, Straussler, & Scheinker.
• It occurs at an estimated annual incidence of 5 cases/100 million
population.
• Neurologic signs and symptoms: cerebellar ataxia, gait
abnormalities, dementia.
• GSS is considered a variant of the familial form of CJD.
Gerstmann Straussler Scheinker 64

Fatal familial insomnia (FFI)
• A familial disease with autosomal dominant inheritance,
have been reported since 1939.
• Patients with severe dementia and bilateral
symmetrical degeneration of the thalamus.
• Progressive insomnia and autonomic dysfunction,
followed by dysarthria, tremor and myoclonus.
• Typical symptom- insomnia
-mutated prions accumulate in the thalamus, with the
result that the patients are unable to sleep.
• Proliferation of astrocytes, support cell of brain.
65

58

DIAGNOSIS
59
 At present, there are no laboratory tests available to
diagnose BSE in the live animal.
 The BSE testing protocol for an initial rapid test called an
Enzyme-Linked Immunosorbent Assay (ELISA).
 If the ELISA test is inconclusive, samples are sent for
confirmatory testing to the National Veterinary Services
Laboratories (NVSL, US).
 Immunohistochemistry & Western Blot conducted at
National Animal Disease Center (NADC, US).
 The tests are designed to detect the presence of BSE-specific
abnormal prion protein in the brain tissue.

Post Mortem Diagnosis
 Histopathology of brain tissue
 Vacuolation of Neurons and neuropil
 Spongiform changes in gray matter
 Detection of abnormal prion protein
 Atrophy of cerebral cortex and cerebellum.
 Proliferation and hypertrophy of the astrocytes and
amyloid plaques.
 Differentials Diagnosis
 Nervous ketosis,
 Hypomagnesaemia,
 Listeriosis,
 Polioencephalomalacia,
 Rabies,
 Brain tumor,
 Lead poisoning
 Spinal cord trauma.
60
(Grassi, et.al., 2016) 42
vacuoles

 All are based on antibodies to detect
prion protein in tissue.
 Immunohistochemistry (IHC)
considered the gold standard.
 Internationally recognized.
 Expensive, required time and
expertise.
 Rapid diagnostic tests
 Western blotting, ELISA.
61

 June 24, 2005
 New BSE confirmatory testing protocol
 ID-Lelystad a new post-mortem test has been developed
that enables screening brain samples for PrP(BSE) within
1 day.
 This BSE test is especially suited for slaughter line
monitoring.
 A preliminary validation study has shown that both
sensitivity and specificity are 100% compared to the gold
diagnostic standard of histopathology.
 IHC & Western Blot
 Confirmatory tests
 Performed with “inconclusive” BSE rapid screening test
results
 Positive result on either test considered positive for BSE
62

VACCINATION/PREVENTION
63
 No effective vaccines available
 Ban on meat and bone meal feeding from endemic
countries.
 Surveillance measures
 Blood/plasma donation restrictions
 There is no treatment for BSE
 Palliative care

64
(Claudia, 2001)

65
Immunohistochemistry
Presence of rounded deposits of
abnormal protein surrounded by
vacuoles, called florid plaques
74
Vacuolar degeneration and spongiosis

PREVENTION & CONTROL
 No Treatment & Vaccine are available
 Import restrictions on live ruminants and ruminant products
from countries known to have BSE endemic.
 Import restrictions expanded to include all European countries.
 FDA “animal feed rule” ƒ
 Banned most mammalian proteins as food source for ruminants
 Scrapie Free Flock Certification Programme
 Surveillance for CJD /BSE
 Blood/plasma donation restrictions.
66

CONCLUSIONS
 The infectious prion proteins are causative agents of many
mammalian TSEs.
 Prion diseases are characterized by deposition of PrPSC.
 No reagent allowing non-invasive, pre-mortem diagnosis of
prion disease.
 The prion have the unique property of being inheritable,
transmissible and infectious in nature.
 Impose import restrictions on live ruminants and ruminant
products from countries known to have BSE endemic.
 Ban the most mammalian proteins as food source for
ruminants feed.
 Strategic policies for monitoring and surveillance of disease
should be implemented.
 Instant need to develop better diagnostic and treatment for
the disease.
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68
Key points

Future studies
 Effective treatment of neurodegenerative disease is
one of the major challenges for biomedical research.
 Clinicians awareness regarding diagnosis and prompt
reporting of prion diseases.
 Develop new method for early diagnosis.
 Precise Epidemiological studies regarding molecular
classification, genetic typing and host strain typing.
69