These notes are exclusively

for Virology Lectures, 3rd Year Medical Students,Dept. Of

Pathology, Egerton University and 2nd year Medical Students
in the Department of Parasitology, Beijing Medical School -2009-
2012

DR M.Abdille, PhD
VIROLOGY

Principles of Virus Architecture

(and a little bit of history)

Design of the protein shell

The complex arrangements of macromolecules in the virus shell are minute marvels of
molecular architecture. Specific requirements of each type of virus have resulted in a
fascinating apparent diversity of organization and geometrical design. Nevertheless, there
are certain common features and general principles of architecture that apply to all
viruses.

In 1956, Crick and Watson proposed on theoretical considerations and on the basis of
rather flimsy experimental evidence then available, principles of virus structure that have
been amply confirmed and universally accepted.
 They first pointed out that the nucleic
acid in small virions was probably
insufficient to code for more than a few
sorts of protein molecules of limited size.
The only reasonable way to build a
protein shell, therefore, was to use the
same type of molecule over and over
again, hence their theory of identical
subunits.

The second part of their proposal concerned the way in which the subunits must be
packed in the protein shell or capsid. On general grounds it was expected that subunits
would be packed so as to provide each with an identical environment. This is possible
only if they are packed symmetrically. Crick and Watson pointed out that the only way
to provide each subunit with an identical environment was by packing them to fit some
form of CUBIC SYMMETRY. A body with cubic symmetry possesses a number of
axes about which it may be rotated to give a number of identical appearances. These
predictions were soon confirmed and it became evident that the occurrence of icosahedral

features in quite unrelated viruses was not a matter of chance selection but that
icosahedral symmetry is preferred in virus structure.

An ICOSAHEDRON

is composed of 20 facets, each an equilateral triangle, and 12 vertices, and because of the
axes of rotational symmetry is said to have
5:3:2 symmetry

Axes of Symmetry

There are, in fact, six 5-fold axes of symmetry passing through the vertices, ten 3-fold
axes extending through each face and fifteen 2-fold axes passing through the edges of an

icosahedron.

Icosahedral symmetry requires definite numbers of structure units to complete a shell. In

their discussions, Crick and Watson (1956), thinking in terms of asymmetrical protein
subunits packed in such a way that each has an identical environment, pointed out that a
virus with 5:3:2 symmetry required a multiple of 60 subunits to cover the surface
completely. Each unit would be related identically and asymmetrically with its
neighbours, and none of the units would coincide with an axis of symmetry.
 The introduction of NEGATIVE STAINING (Brenner and Horne, 1959)
revolutionized the field of electron microscopy of viruses. Within just a few
years, much new and exciting information about the architecture of virus
particles was acquired. Not only were the overall shapes of particles revealed
but also the symmetrical arrangement of their components. This led to a need
for a new terminology to describe the viral components.

Lwoff, Anderson and Jacob (1959) proposed the terms "capsid" and "capsomers" to
represent, respectively, the protein shell and the units comprising it, and the
term"virion" to denote the complete infective virus particle (i.e. a capsid enclosing the
nucleic acid). This terminology was generally accepted although it later proved to be
inadequate.

As soon as the first high resolution micrographs of negatively stained icosahedral viruses
were obtained (Horne et al., 1959 - adenovirus; and Huxley and Zubay, 1960 - turnip
yellow mosaic virus) it seemed that there was a structural paradox. The number of
morphological units observed on the surface of known icosahedral viruses at that time
was never 60 or multiples of 60, and was often more than 60. Furthermore, the capsomers
themselves appeared to be symmetrical and were located on symmetry axes, e.g.
herpesvirus.

Below is a model of the herpes simplex virus capsid

There was direct evidence that capsomers of herpesvirus were hexagonal and pentagonal
in section. It is obvious that five-fold capsomers must be located on axes of five-fold
symmetry, and six-fold capsomers may be situated on axes of two-fold or three-fold
symmetry, or in indifferent sites where they are suited to hexagonal packing.

It was therefore clear that the capsomers were not equivalent to the subunits of Crick and
Watson (1956).
An obvious solution to the problem was provided by supposing that the symmetrical
capsomers are built from a number of ASYMMETRICAL SUBUNITS. In this way it is
possible to build a variety of complicated bodies in which 5:3:2 symmetry is preserved
and in which the number of subunits is a multiple of 60 as predicted by Crick and
Watson.

The theoretical basis for the structure of isometric viruses was put on a firm foundation
by Caspar and Klug (1962) with their concept of identical elements in quasi-equivalent
environments. They defined all possible polyhedra in terms of structure units. The
icosahedron itself has 20 equilateral triangular facets and therefore 20T structure units
where T is the TRIANGULATION NUMBER given by the rule:
 T=Pf where P can be any number of the series 1,3,7,13,19,21,31 ..(=h + hK +K , for
all pairs of integers, h and K having no common factor) and f is any integer.

Morphological units can be clustered as 20T trimers, 30T dimers or separated as 60T
monomers. The number of morphological units that would be produced by a clustering
into hexamers and pentamers can be calculated as follows: There are 10(T-1) hexamers
plus 12. (and only 12) pentamers.

Caspar and Klug (1962) claimed that most icosahedral viruses fall into2 classes:- P=1
and P=3; and that all deltahedra for which P=>7 are skew, and therefore exist in right and
left- handed forms. One "hand" might be selected by the nucleic acid, but there would
still be the chance that mistakes in assembly leading to defective particles might occur.
The most probable mistake in assembly would be the formation of tubular forms. Tubular
structures which have a diameter and surface structure similar to icosahedral virus
particles have been observed associated with polyoma and papilloma viruses.

In a review of symmetry in virus architecture, Horne and Wildy (1961) showed that all
the viruses then known (with the exception of a few bacteriophages) fell into two main
morphological groups:-
those with cubic symmetry
and the others with HELICAL symmetry.
"Linear" viral capsids have RNA genomes that are encased in a helix of identical protein
subunits.
The length of the helical viral nucleocapsid is determined by the length of the nucleic
acid.

Until 1960, the only known examples of virions with helical symmetry were those of
plant viruses,
the best studied example being tobacco mosaic virus.
At that time, the architecture of the myxoviruses was poorly understood. Early electron
micrographs of shadow-cast preparations revealed particles of varying shape and size but
little detail could be reported (Bang, 1948). With the advent of negative staining, it
became obvious that the myxo- and paramyxo-viruses consisted of an inner nucleo-
protein component with helical symmetry surrounded by an envelope of characteristic
morphology. This realization of the helical symmetry of the myxoviruses laid the
foundation for the understanding of the symmetry of other complex groups of viruses
such as rabies virus and granulosis virus.

In an attempt to clarify the terminology for virus components, Caspar et al. (1962)
made a number of proposals which were generally accepted. Briefly, the proposals are as
follows:

1. The CAPSID denotes the protein shell that encloses the nucleic acid. It is built of
structure units.
2. STRUCTURE UNITS are the smallest functional equivalent building units of the
capsid.
3. CAPSOMERS are morphological units seen on the surface of particles and
represent clusters of structure units.
4. The capsid together with its enclosed nucleic acid is called the
NUCLEOCAPSID.
5. The nucleocapsid may be invested in an ENVELOPE which may contain
material of host cell as well as viral origin.
6. The VIRION is the complete infective virus particle
 To provide contrast in the
electron microscope

Visualization of individual virus particles

Most biological materials show little contrast with their surroundings unless they are
stained. In the case of light microscopy, contrast can be enhanced by using coloured
stains which selectively absorb certain wavelengths. The electrons in the electron
microscope are absorbed very little by biological material and contrast is obtained mainly
by electron scattering.

To heighten the contrast between viruses and the background, use is made of electron-
dense "stains". These are usually compounds of heavy metals of high atomic number, that
serve to scatter the electrons from regions covered with the stain. If virus particles are
coated with stain (positive staining), fine detail may be obscured. Negative staining
overcomes this problem by staining the background
and leaving the virus relatively untouched. The
negative stain is moulded round the virus particle,
outlining its structure, and is also able to penetrate
between small surface projections and to delineate
them. If there are cavities within the virus particle
that are accessible to the stain, these will be revealed
and some of the internal structure of the virus may
be disclosed.

For negative staining, use is made of transmission

electron microscopy, and only electrons which
pass through the specimen are involved in the
formation of the final image.
When particles are covered with stain, components
of both top and bottom surfaces of a three-dimensional structure are contained in the
two-dimensional image. This superposition can make it difficult to distinguish fine
structures which would ordinarily be well within the resolution of the electron
microscope.
 One-sided staining provides more accurate information about the
organization of capsomers, etc., although these particles are less
well supported and tend to "collapse", resulting in apparent
increase in size.

When two surfaces of regularly repeating units (such as the surface

of some virus particles) are superimposed slightly out of register,
moire pattern artefacts may result which could lead to false interpretation of particle
ultrastructure.

Below is an example of rotaviruses stained from below, and from both

sides (top and bottom)

Rotaviruses have a lattice-like arrangement of capsomers (left), and the large ring shapes
(right) are artefacts produced by moire effects.

Superposition artefacts are seen in this semi-crystalline array of an isometric virus from
the insect Nudaurelia.

If you can't spot the "ban-the-bomb" sign, it probably means that you have youth on your
side!
 ENTERO-
VIRUSES

INTRODUCTION

These are small non-enveloped isometric viruses that multiply in the gut
mucosa and are transmitted from person to person by the faecal-oral route
(ingestion disease). They are spread throughout the body via the blood stream. Most
infections occur during childhood, and they are usually transient but produce lifelong
immunity. Clinical syndromes are generally mild, but occasional infections may cause
serious disease e.g. paralytic poliomyelitis, meningitis, or myocarditis. There is a high
degree of serological cross reactivity between the 72 members.

CLASSIFICATION

Viruses belong to the family Picornaviridae (pico=SMALL - RNA viruses)

Enteroviruses:
Polio 1, 2, 3
Coxsackie A 1-24
Coxsackie B 1-6
ECHO 1-34
Entero 68-71
Entero 72 (Hepatitis A)

Rhino viruses: > 120 serotypes

Other animal viruses: e.g. Foot & Mouth Disease virus

POLIOMYELITIS

Poliovirus has been well studied and is a good example of an enterovirus.

Virus: small (30nm) and stable; an icosahedral capsid enclosing a positive-sense, single-
stranded RNA genome. Relatively resistant to extremes of pH and temperature, and to
lipid solvents and detergents.

Types: 3 types can be distinguished by antigenic properties.

CLINICAL

Source: Only known source is infected man

Incubation: After ingestion of the virus, there is local multiplication in the oropharynx
and associated lymph nodes. Local multiplication also takes place in the gut mucosa and
regional lymph nodes. Thereafter a viraemia follows, and the patient may experience a
fever about a week after exposure.

Illness: Most infections are asymptomatic, although in some there is a minor transient
febrile illness. Occasionally (between 1/100 and 1/1000 of cases) the viraemia may lead
to CNS involvement and paralysis due to permanent damage to the anterior horn motor
neurones of the spinal cord. The patient may experience degrees of headache, fever,
meningism, aseptic meningitis and muscle pains, and finally muscle paralysis, usually
asymmetrical. Paralysis develops more frequently in adults, and may be precipitated by
muscle trauma (injections, exercise), tonsillectomy, pregnancy and steroid drug
administration. The spinal cord may be damaged in a progressive manner from distal to
more central - some cases may progress to involve the medulla and brainstem (bulbar
paralysis) with consequent respiratory paralysis and death, or life on a respirator.

Virus is produced and released into the gut (and throat initially) and can be isolated from
the throat or stools for some weeks following the incubation period. No true long term
carrier status occurs.

The host's antibody response begins soon after the viraemia. Good solid lifelong
immunity results to the specific strain of poliovirus, but subsequent infection with other
strains may still occur.

DIAGNOSIS

(1) Demonstration of the virus

Virus may be recovered from faeces (also throat swabs), by inoculation of cell cultures
and recognition of cytopathic effects with confirmation by neutralisation of infectivity
with specific antisera. Vaccine strains may be recovered and need to be differentiated
from wild strains by molecular nucleic acid techniques (PCR). Multiple specimens over
several days improves chances of recovery of the virus.
(2) Serology:
Most cases of poliomyelitis that come to medical attention present with paralysis, i.e.
quite late in the pathogenesis, and antibodies have already been formed. Antibodies are
not usually helpful in providing a positive diagnosis of poliomyelitis, but do give the
immune status of an individual (does/does not need further vaccination). Detection of
specific IgM has not been applied to polio diagnosis. Antibodies are traditionally tested
by micro-neutralisation of infectivity in vitro using antisera to known virus strains

CSF: Polio virus is never found in the CSF but antibodies here mean either CNS infection
or a leak from blood antibodies.

EPIDEMIOLOGY

Before the introduction of a vaccine, (< 1960), polio was endemic in the tropics, with
rapid circulation in young children (poor hygiene facilitates faecal-oral spread) with
minimal paralysis (? protective effect of residual maternal antibody). This ensured high
"herd immunity" without epidemics.

In temperate regions, polio showed peaks in summer/autumn. As conditions of hygiene

improved, viral spread was hindered and the age of primary exposure rose. Primary
infection in adulthood resulted in a much higher incidence of paralytic disease. These
tendencies rose to a climax in Scandinavian countries just before the vaccine era ,when
they experienced devastating epidemics of paralytic poliomyelitis.

Post Vaccine. Universal vaccination commencing in the early 1960s, and has eliminated
polio from the Western world, including North, Central and South America. The disease
has also largely been controlled it in Africa and Asia. In South Africa, polio has been
effectively eliminated but reintroductions into regions with deficient vaccination
programs has resulted in localised outbreaks (e.g. 1982 and 1988).
The World Health Organisation is forging ahead with a total elimination plan (c.f.
smallpox) "by the year 2000".

POLIO VACCINES

(1) Live attenuated virus (SABIN)(1963)

Strains of poliovirus 1, 2 and 3 which have been attenuated by passage in unnatural

conditions to lose neurovirulence.
3 live strains mixed, given as oral drops (easy administration).
Given on 3 occasions plus boosters (one strain may interfere with uptake of another,
hence must be given repeatedly to ensure immunity to all 3 types).
Wild enteroviruses coincidentally present in the gut may also interfere, especially in the
tropics.

Live vaccination mimics natural infection with good immunity including IgA in the gut.
This vaccine is used in RSA, USA and most other countries. It is very important to
maintain 'cold chain' when storing and distributing vaccine as it may lose potency.

(2) Killed whole virus (SALK)(1957)

Polio 1, 2 and 3 grown in cell cultures, mixed, killed with formalin.

3 injections at 3 to 6 months of age; later boosters.
Much antigen is required which makes the vaccine expensive.
Effective when coverage is good (nearly 100% immunity in people regularly vaccinated).
Still used in some countries (Netherlands, Scandinavia) in 1990's.

SABIN POLIO VACCINE POLICY, RSA( 1989 Onwards)

A. Neonatal monovalent P1 at birth

B. Standard Trivalent (P1 + P2 + P3) at 3 months, 4½ months and 6 months of age

C. Boosters: at 18 months, and again at time of School entry.

Polio is controlled (?eliminated) by:

(1) Education
(2) Vaccination
(3) Surveillance

ENTEROVIRUSES - OTHER THAN POLIO

i.e. Coxsackie, Echo, Entero 68-72

Virus structure, Epidemiology, Pathogenesis of all the enteroviruses are remarkably

similar and follow the pattern described for polio.
Most infections are silent. Viraemia may lead to degrees of involvement of secondary
'target organs' and clinical symptoms and signs related to those organs. For example, the
most common type of meningitis seen in the Cape is aseptic meningitis caused by
coxsackie or echo viruses (which can often easily be isolated from the CSF, in contrast to
polio). Viral meningitis resolves spontaneously without treatment but bacterial meningitis
is a medical emergency requiring treatment.

Enteroviruses may be found in the gut of healthy as well as sick children; the association
with any illness may be purely co-incidental.
 CLINICAL SYNDROMES

Information given in the following format:-

CLINICAL SYNDROME
Common virus associated with the clinical syndrome
Less commonly associated virus

ASYMPTOMATIC
All enteroviruses

PARALYSIS - permanent

Polio 1, 2, 3
Coxsackie A7

PARALYSIS - temporary
Coxsackie B1-6

MENINGITIS (aseptic)Echo, Coxsackie A and B

Polio, Entero 71

ENCEPHALITIS
Entero 71
Polio, Echo

RASH
- macular
Many enteroviruses
- vesicular - (e.g. 'Hand Foot Mouth')
Coxsackie A

SUMMER FEBRILE ILLNESS

Many enteroviruses

VESICULAR PHARYNGITIS ('Herpangina')

Coxsackie A

MYOCARDITIS
Coxsackie B

EPIDEMIC MYALGIA ('Bornholm')

Coxsackie B
UPPER RESPIRATORY INFECTION (cold)
Echo, Coxsackie A

PANCREATITIS
Coxsackie B

GASTRO-ENTERITIS
Many enteroviusess

CONJUNCTIVITIS (Haemorrhagic)
Entero 70

HEPATITIS
Entero 72 (hepatitis A virus)
 VIRAL
GASTRO-
ENTERITIS

INTRODUCTION

Paediatric diarrhoea remains one of the major causes of death in young children. This is
especially so in Asia, Africa and Latin America where it causes millions of deaths in the
age group 0-4 years.

The main factors for high incidence and mortality are unsafe water or inadequate
sanitation, requiring social, economic and political solutions. The immediate causes are
often of an infectious nature and include a variety of pathogenic micro-organisms. A
range of bacteria and parasites has been identified = enterotoxigenic Escheritia coli,
salmonella, shigella, cholera, other vibrio bacteria, as well as cryptosporidium, but these
account for well below half of investigated cases.

A number of different viruses cause diarrhoea, of which the most important is the
family of ROTAVIRUSES.

Rotaviruses have been estimated to cause 30-50% of all cases of severe diarrhoeal
disease in man.

Two strains of adenovirus (40 and 41) have also been associated with diarrhoeal disease.

A group of "small round viruses" (discovered by electron microscopy) have been

linked by genetic techniques as closely related to the previously described "Norwalk"
agent, associated with vomiting and diarrhoea.

ROTAVIRUS - (REO virus family)

• Group A subtypes 1, 2, 3, 4 (main human pathogens)

(Further 7 subtypes) also infect animals (monkey, calf, mouse)
• Group B Infects pigs and rats
Found to cause extensive outbreaks in China in past decade
 • Group C Infects Pigs (Occasionally Man)
• Group D Infects birds
• Group E Infects pigs

VIRUS MORPHOLOGY:
Particles are 70 nm round, double shelled, enclosing a genome of 11 segments of double
stranded RNA.
The virus is hardy and may even survive in sewage, despite stringent treatment.
Human rota virus has proved difficult to culture in vitro, but the serologically related
monkey and calf rotaviruses grow easily in cell culture.

CLINICAL:
Essentially an ingestion disease (faecal-oral route)

Incubation is short : 1 to 3 days.

Illness: Sudden onset watery diarrhoea, with or without vomiting. May last up to 6 days
(or longer if immunocompromised). The disease is self limiting.
Complications: Dehydration may result, this can be severe and life threatening in young
children.
Treatment: No specific treatment of viral infection is available nor is it really required.
Treatment is aimed at prevention and/or treatment of dehydration by oral and/or
intravenous fluids and electrolytes
Diagnosis: Detection of virus in stools (peaks at day 3 or 4 of diarrhoea):-

1. Latex agglutination
2. Elisa
3. Electron Microscopy (labour intensive, relatively insensitive)
4. Electrophoresis of RNA segments

(Antibody can be detected but is not clinically useful)

Prevention:
Non specific factors: improved hygiene, education, clean water
Specific - Breast feeding helps to provide passive immunity in the newborn (from
maternal antibodies),
Vaccination is still experimental.

EPIDEMIOLOGY
Infection is found world-wide.
All ages can be infected and reinfection can occur (usually asymptomatic).

Age: Infections at < 6 months age and > 5 years of age tend to be asymptomatic and
give degrees of protection against diarrhoeal infection.
Maternity hospitals commonly have resident strains which readily cause asymptomatic
infections of new- borns.
Seasons: In temperate '1st world' populations rota virus is the main cause of winter
gastroenteritis .
In tropical and developing countries, rotavirus diarrhoea occurs all the year round ,but
with a peak in summer. However, it is only one of a variety of pathogens causing
diarrhoea.
Vaccine: In view of the major role of dehydration from diarrhoea as a cause of childhood
death, the World Health Organisation has waged an intensive campaign for

• (1) oral rehydration solutions to prevent or treat dehydration

• (2) development of a vaccine for rota virus infection.

No vaccine is currently used routinely, but several candidate vaccines are being evaluated
in children: e.g. animal strains, attenuated human strains, animal-human recombinant
strains, designed to cover all 4 main human pathogenic strains.
The prevention of severe dehydration is the main aim, rather than totally preventing
infection.

ADENOVIRUS

A limited number of strains of ADENOVIRUS have been causally related to childhood

diarrhoea.
They do not grow in cell cultures and were discovered by Electron Microscopy. (
Recently there has been limited success in special cell culture systems).
They are classified in the 40/41 serogroup of adenoviruses.
Viruses can be isolated from stools,as well as throat and respiratory secretions.
Adenoviruses in stools can be detected by latex agglutination, and the 40/41 strains can
be detected by specific molecular techniques.
The exact role or significance of these strains in the global picture of childhood
diarrhoea, especially in developing countries, is not yet fully established.

Apart from this 40/41 group, other adenoviruses may be found in the stools of
asymptomatic children. .

GASTROENTERITIS IN OLDER CHILDREN AND ADULTS

Apart from the severe problem of diarrhoea in young children there have been outbreaks
of infectious gastro-enteritis in adults. Electron microscopy (of the stools) has revealed
two main groups of virus particles which do not grow in cell culture.

• (1) Calici viruses (ssRNA) including Norwalk and related agents ('Hawai';
Ditchling; 'W')
• (2)"small round viruses" about which very little is known

NORWALK agent (33nm in size)

'Common source' type of explosive outbreaks of gastroenteritis, with limited secondary

spread to household contacts, have been described. These often occur in institutions, or
follow common source ingestion episodes e.g. celebratory feasts.
Vomiting with cramps are more common symptoms than the diarrhoea.

At first, the virus was seldom identified as there were no easy diagnostic tests - only
expensive electron microscopy of stools. Serology was limited, as the only antigen
available was prepared from known infected stools - not in plentiful supply!

Recently, molecular techniques have shown that many of these agents from different
parts of the world are essentially similar. Molecular techniques have also enabled the
expression of viral antigens that can be used in serological surveys. For example, a 1993
survey in the UK has shown that Norwalk infection is apparently a common silent
infection in childhood. Antibody prevalence rises to about 16 years of age and then
begins to level off, with 80% of persons above 30 years being seropositive. It was
hitherto believed that Norwalk infections were rare, - it now seems that the disease is
rare, not the infection.
 VIRAL HEPATITIS
The termVIRAL HEPATITIS is usually used to describe infections
caused by agents whose primary tissue tropism is the liver.
To date, at least five hepatitis viruses have been recognised, and these
have been named:-
Hepatitis A, B, C, D and E.
Acute hepatitis may also occur as part of the clinical course of a number
of viral infections, including
human cytomegalovirus, Epstein-Barr virus, herpes simplex virus, yellow fever
virus and rubella.

Clinical Features

Hepatitis due to all these viruses presents clinically in a very similar fashion, especially
during the acute phase of the illness. Thus a specific diagnosis can only be made in the
laboratory.
The majority of infections are totally asymptomatic, but common clinical features
include: anorexia, nausea, vomiting, right upper quadrant pain and raised liver enzymes
AST and ALT.
Jaundice is the hall mark of infection, but tends to develop late.
Anicteric cases are also very common.

ENTERICALLY TRANSMITTED HEPATITIS: A and E

PARENTERALLY TRANSMITTED HEPATITIS B , C , D and G

Hepatitis A - "Infectious Hepatitis"

Caused by a picornavirus, Enterovirus 72

This is a small, non-enveloped icosahedral particle, 27 nm in diameter, containing a
ssRNA genome
 Clinical Features
Incubation period 3-5 weeks (mean 28 days)
Milder disease than Hepatitis B; asymptomatic
infections are very common, especially in
children.
Adults, especially pregnant women, may
develop more severe disease.
Although convalescence may be prolonged,
there is no chronic form of the disease.

Complications:
Fulminant hepatitis is rare: 0.1% of cases

Pathogenesis
Virus enters via the gut; replicates in the alimentary tract and spreads to infect the liver,
where it multiplies in hepatocytes.
Viraemia is transient. Virus is excreted in the stools for two weeks preceding the onset
of symptoms.

Epidemiology
World-wide distribution; endemic in most countries. The incidence in first world
countries is declining. There is an especially high incidence in developing countries and
rural areas.
In rural areas of South Africa , the seroprevalence is 100%.

Transmission - Enteric
Large numbers of virus particles are excreted in stools, before the onset of symptoms.

1) Case-to-case, via faecal-oral route.

Outbreaks in creches are very common.
2) Contamination of food or water with sewage
Infected food handlers
Shell fish grown in sewage-polluted water

Diagnosis
Virus cannot be cultured in vitro from clinical material, and diagnosis is made on the
presence of HAV-specific IgM in the patient's blood.

Prevention
1) Passive immunisation -

Normal immunoglobulin given to:

Travellers to third world countries
Household contacts of acute cases
2) Active Immunization
Inactivated cell culture-derived vaccine has recently become available; not in general use

Hepatitis E

Recently identified cause of enterically transmitted non-A, non-B (NANB) hepatitis

Calicivirus
spherical, non enveloped, 27-34 nm particles containing a ssRNA genome.

Clinical Features
Incubation period 30-40 days
Acute, self limiting hepatitis, no chronic carrier state
Age: predominantly young adults, 15-40 years

Complications
Fulminant hepatitis in pregnant women. Mortality rate is high (up to 40%).

Pathogenesis
Similar to hepatitis A; virus replicates in the gut initially, before invading the liver, and
virus is shed in the stool prior to the onset of symptoms.
Viraemia is transient. A large inoculum of virus is needed to establish infection.

Epidemiology
Little is known yet. The incidence of infection appears to be low in first world countries.

1) Large outbreaks have been described in India, Mexico and North Africa where the
source of infection is usually gross faecal contamination of drinking water supplies.
2) Case-to-case transmission to household contacts appears to be uncommon. This
suggests that a large inoculum is needed to establish infection.

The incidence of infection in South Africa is unknown.

Diagnosis
No routine laboratory tests are available as yet. Virus cannot be cultured in vitro.

1) Calicivirus-like particles in the stool, by electron microscopy

2) Specific IgM in serum
3) PCR HEV-specific sequences in stool
 PARENTERALLY TRANSMITTED HEPATITIS
B, C, D and G

Hepatitis B

Hepadna virus
42nm Virions (also known as "Dane particles") contain a circular dsDNA genome

HBV Antigens
HBsAg = surface (coat) protein
produced in excess as small spheres and tubules
HBcAg = inner core protein
HBeAg = secreted protein; function unknown

You can view a diagram and electron micrographs of the virus structure.

Clinical Features
Incubation period 2 - 5 months
Insidious onset of symptoms. Tends to cause a more severe disease than Hepatitis A.
Asymptomatic infections occur frequently.

Pathogenesis
Infection is parenterally transmitted. The virus replicates in the liver and virus
particles, as well as excess viral surface protein, are shed in large amounts into the blood.
Viraemia is prolonged and the blood of infected individuals is highly infectious.

Complications
1) Persistant infection:-
Following acute infection, approximately 5% of infected individuals fail to eliminate the
virus completely and become persistantly infected.

Those who are at particular risk include:

babies, young children
immunocompromised patients
males > females

The virus persists in the hepatocytes and on-going liver damage occurs because of the
host immune response against the infected liver cells.

Chronic infection may take one of two forms:

Chronic persistent Hepatitis - the virus persists, but there is minimal liver damage
Chronic Active Hepatitis - There is aggressive destruction of liver tissue and rapid
progression to cirrhosis or liver failure.
2) Patients who become persistently infected are at risk of developing hepatocellular
carcinoma (HCC)..
HBV is thought to play a role in the development of this malignancy because:

a) 80% of patients with HCC are carriers of hepatitis B.

b) Virus DNA can be identified in hepatocellular carcinoma cells.
c) Virus DNA can integrate into the host chromosome.

3) Fulminant Hepatitis
Rare; accounts for 1% of infections.

Epidemiology

Prevalence of disease in Africa

World-wide there are 450 million persistant carriers of hepatitis B, 50 million of which
are in Africa. Carriage rates vary markedly in different areas. In South Africa, infection is
much more common in rural communities than in the cities. Hepatitis B is parenterally
transmitted

1) Blood:

• Blood transfusions, serum products,

• sharing of needles, razors
• Tattooing, acupuncture
• Renal dialysis
• Organ donation

2) Sexual intercourse

3) Horizontal transmission in children, families, 'close personal contact'.

This is the major mode of transmission in South Africa where the majority of individuals
become infected at between three and nine years of age.
Horizontal transmission also occurs in children's institutions and mental homes.

4) Vertical transmission - perinatal transmission from a carrier mother to her baby

• transplacental (rare)
• during delivery
• post natal , ??breast feeding , ??close contact

( This is the major mode of transmission in South East Asia)

Diagnosis: Serology

A. Acute infection with resolution

 Viral antigens:
1) Surface antigen (HBsAg) is secreted in
excess into the blood as 22 nm spheres and
tubules. Its presence in serum indicates
that virus replication is occurring in the
liver
2) 'e' antigen (HBeAg) secreted protein is
shed in small amounts into the blood. Its
presence in serum indicates that a high
level of viral replication is occurring in the
liver
3) core antigen (HBcAg) core protein is
not found in blood

Antibody response:
1) Surface antibody (anti-HBs) becomes detectable late in convalescence, and indicates
immunity following infection. It remains detectable for life and is not found in chronic
carriers (see below).
2) e antibody (anti-HBe) becomes detectable as viral replication falls. It indicates low
infectivity in a carrier.
3) Core IgM rises early in infection and indicates recent infection
4) Core IgG rises soon after IgM, and remains present for life in both chronic carriers as
well as those who clear the infection. Its presence indicates exposure to HBV.

B. Serology of the chronic carrier

Prevention

1) Active Immunization

Two types of vaccine are available:

Serum derived - prepared from HBsAg purified from the serum of HBV carriers
Recombinant HBsAg - made by genetic engineering in yeasts

Both vaccines are equally safe and effective. The administration of three doses induces
protective levels of antibodies in 95% of vaccine recipients.
Universal immunization of infants was introduced in April 1995. Infants receive 3 doses
at 6, 10 and 14 weeks of age.

Vaccine should be administered to people at high risk of infection with HBV:

1) Health care workers
2) Sexual partners of chronic carriers
3) Infants of HBV carrier mothers

2) Passive Antibody
Hepatitis B immune globulin should be administered to non immune individuals
following single episode exposure to HBV-infected blood.
For example: needlestick injuries

Hepatitis C

The major cause of parenterally transmitted non A non B hepatitis. It eluded

identification for many years. In 1989, the genome was cloned from the serum of an
infected chimpanzee.

Virology

Putative Togavirus related to the Flavi and Pesti viruses.

Thus probably enveloped.
Has a ssRNA genome
Does not grow in cell culture, but can infect Chimpanzees

Clinical Features
Incubation period 6-8 weeks
Causes a milder form of acute hepatitis than does hepatitis B
But 50% individuals develop chronic infection, following exposure.

Complications
1) Chronic liver disease
2) Hepatocellular carcinoma

Epidemiology

Incidence endemic world-wide; high incidence in Japan, Italy and Spain

In South Africa, 1% blood donors have antibodies

Transmission

• Blood transfusions, blood products

• organ donation
• Intravenous drug abusers
• community acquired: mechanism unclear. ?Vertical transmission
• ?sexual intercourse

Diagnosis

1) Serology
Reliable serological tests have only recently become available.
HCV-specific IgG indicates exposure, not infectivity

2) PCR detects viral genome in patient's serum

Delta Agent
Defective virus which requires Hepatitis B as a helper virus in order to replicate.
Infection therefore only occurs in patients who are already infected with Hepatitis B.

Clinial Features
Increased severity of liver disease in Hepatitis B carriers.

Virology
virus particle 36 nm in diameter
encapsulated with HBsAg, derived from HBV
delta antigen is associated with virus particles
ssRNA genome
Epidemiology
Identified in intra-venous drug abusers in Italy. Incidence in South Africa is unknown.

Hepatitis G (HGV)

A virus originally cloned from the serum of a surgeon with non-A, non-B, non-C
hepatitis, has been called Hepatitis G virus. It was implicated as a cause of parenterally
transmitted hepatitis, but is no longer believed to be a major agent of liver disease. It has
been classified as a Flavivirus and is distantly related to HCV.

ARBOVIRUSES
and viral
HAEMORRHAGIC FEVERS

GROUP COMMON FEATURES

There is a large heterogenous group of RNA viruses (>400), which cause essentially
zoonotic infections of wild animals and are spread by insects (hence the name
arboviruses) and occasionally cause problems in man. They have the unusual ability to
multiply in the vertebrate host (usually mammals) as well as in the very different
metabolism of the insect vector.

DISTRIBUTION
They are mostly found in the warmer parts of the world (tropics) but some range into
temperate regions. Rapid transport today means every country is at some risk.

NATURE
The majority are zoonoses maintained silently in nature by their animal (vertebrate) hosts
which form their natural reservoir. In their normal hosts they have usually adapted well
and do not produce overt disease. They produce high blood titres of virus which favours
their chances of being picked up by a blood-sucking insect vector. Within that vector they
can multiply again, which favours their chances of being transmitted from the insect
vector to a new member of their definitive animal host. Crossing into non-normal hosts
may produce disease.

HUMAN INFECTION
Man usually forms an accidental, non-normal host, with sporadic - end point’ infections
which have no further transmission. However, there are some notable exceptions to this
generalisation, where human infection and disease can be spread to other humans via
human-feeding insects or even directly person to person. Human infections thus tend to
be isolated or sporadic, unless climatic conditions have favoured large-scale vector
proliferation when human (and animal) infections may become seasonal and epidemic.
Some vaccines but no specific treatments are available.

TRANSMISSION
Climate has a major role in facilitating transmission, eq. mosquito proliferation after
heavy (warm) rains. Viral replication only occurs in the insect vector in warm conditions
(>25’C). Human activity can bring man into contact with an otherwise silent cycle in
nature, eg. clearing of forests, agriculture. Creating vector promoting activities, such as
dams and irrigation, may enhance human arbovirus disease potential. Within the vector
the virus multiplies without making the insect sick and in some viruses there is evidence
there may be trans-ovarial (vertical) spread, ie. the virus can overwinter from one season
to the next inside the insect eggs. The virus is present in the insect saliva and the infection
is lifelong in the insect.

LABORATORY
Arboviruses are single-stranded enveloped RNA viruses with haemagglutinating
properties. They grow in suckling mice and/or in cell cultures. Mouse inoculation is
hazardous to human workers and must only be done in special containment facilities
(National Institute for Virology in RSA). Diagnosis is made by isolation of the
virus (usually done in a specialist laboratory) and by detection of IgM and IgG
antibodies by haemagglutination-inhibition, or ELISA.

CONTROL
Human disease control is limited to prevention by

1. control of vector, eg. mosquito control which con be very effective; and,
2. use of vaccines in a few types of infection, eg. Yellow Fever,
3. surveillance.

HUMAN DISEASE

Infection of man may produce a wide spectrum of disease - from subclinical infections to
fulminant haemorrhagic disease with death. By and large, there are 4 major clinical
patterns of disease. Any one virus characteristically tends to produce mostly one pattern,
but overlapping may occur.

Major Clinical Categories of Arboviral Disease

• No clinical illness.
• Febrile systemic illness.
• Encephalitis
• Haemorrhagic Fever.

1) No clinical illness
Severe clinical disease in man attracts attention, and many arboviruses were first
discovered (and still are today) by investigation of unexplained, possibly infectious,
disease cases. As the investigation progressed, it was commonly found that there were
milder disease cases in the vicinity of a severe disease case, aa well as subclinical cases
of infection. We now know that subclinical cases are probably more common than frank
disease in any one outbreak, eg. even in a yellow fever epidemic. New Arboviruses have
also been discovered by intensive cultural and serological investigation of possible insect
vectors (mosquitos, ticks) and possible definitive hosts (rodents, birds, small mammals,
etc.).

2. Febrile systemic illness

After inoculation there is a short incubation period of a few days, and then an abrupt
onset of disease at a time of high viraemia. There is a sudden onset of fever with chills;
headache; muscle, bone and joint pains; nausea and vomiting, and often a rash and
lymphodenopothy. These clinical features may be mild or very severe and prostrating.
The disease is typically self-limiting and short lived (few days to a week). Occasional
cases may go on to a more fulminant form (- see haemorrhagic fever section).

Examples

In South Africa.

West Nile Virus.

Asia, Central and South Africa (Transvaal).
Birds and mosquitoes. Human outbreaks after rains.
Rift Valley Fever.
East Africa, Egypt, Israel, South Africa.
Disease (with abortions) in sheep, cows, goats, spread by mosquitoes.
? occasional direct droplet spread.
Definitive host not identified.
Human outbreak follows animal outbreak after rains.
Sindbis
Africa & SE Asia (Transvaal, OFS)
Birds/mosquitoes.
Mild "Summer popular fever" epidemics in man.

Wesselsbron, Chikungunya

Other than South Africa:

Dengue.
 SE Asia,, Carribean and Central America.
Mosquitoes
Definitive host is unknown (? is none).There are seasonal epidemics in man.
4 Serological strains without cross-protection.
Previous exposure may predispose to haemorrhagic form.
(Has been imported into RSA.)

3. Encephalitis

Almost all arboviruses may infect the CNS but some characteristically produce an
encephalitis in man.
Symptoms start with fever, progressively severe headache, nausea and vomifing, neck
stiffness, paralysis, decreased consciousness, convulsions and death in severe cases.

Examples (none in South Africa)

Japanese B Encephalitis

SE Asia - seasonal.
Birds/mosquitoes - transmitted to man.
2 Human vaccines available..
(Closely related Saint Louis Encephalitis - USA, Murray Valley Encephalitis -
Australia)
Eastern Equine Encephalitis. (EEE).
Eastern USA, Central & South America.
Birds/mosquitoes.
Severe CNS disease in horses and man.
Western Equine Encephalitis. (WEE)
North & South America.
Birds/mosquitoes.
Severe disease in horses. Milder disease in man.
Tick Borne Encephalitis (TBE)
Central Europe, East Russia.
Goats/ticks.
Goats milk seems to be infectious.

4. Haemorrhagic Fever

A number of arboviruses (Toga viruses and Bunyaviruses) and some similar viruses
(Arenaviruses and Filoviruses) may cause an illness similar to the rather non-specific flu-
like illness described under (2) above, but which rapidly progresses to a severe
disseminated illness with a marked bleeding tendency and multi-organ failure.

There is usually a considerable mortality in these haemorrhagic fever infections.

 Note:- Overwhelming parasitaemia/septicaemia may produce a similar clinical
picture and require to be excluded,
eg. meningococcus, rickettsia (Tick Bite Fever), leptospirosis, hepatitis, malaria,
snake bite, tryponosomiasis.

Mild and subclinical cases also do occur, but are less easily recognised.

Examples:

In South Africa:

Crimean Congo Haemorrhagic Fever

(Eastern Europe, Central and Southern Africa)

See "What is CCHF?"

A rare disease, but the most common viral haemorrhagic fever in South Africa.
Most frequent in Kimberley area, but antibodies are widespread in cattle herds all over
South Africa.
Transmitted by ticks with striped legs (bont-poot bosluis, Hyalomma species).

Host:
cattle, ticks, ? other small mammals.
History:
visit to country in post week;
contact with animals;
tick bite;
contact with known human case (blood is highly infectious, can spread direcfly).
Illness:
short incubation (sudden onset - headache, fever, chills, body pains; diarrhoea &
vomiting;
dizzyness, confusion, abnormal behaviour,.
pharyngitis, conjunctivitis, red face and neck,
haemorrhage starts day 3 - 6 (not all cases),
multi-organ failure, that leads to death.
Treatment:
isolation and barrier nursing;
transfusions,.
ribavirin.

Outside South Africa:

YellowFever (YF)
Africa:
Central, West East.
Central South America (Amazon basin).
Hosts: Monkey - mosquito (=silent reservoir).
Jungle (sylvan) YF
sporadic cases in forest worker/visitor.
Urban )YF
YF introduced into towns;
now get cycle: man - mosquito - man.
NB. Mosquito control is important.
Illness: as above, but jaundice is a prominent feature (liver necrosis).
Vaccine:
i) "17 D strain" live attenuated virus
Good safe long lasting protection but labile and expensive to administer.
Used in the RSA.
ii) "French Dakar Vaccine" live attenuated brain tissue-derived vaccine
Inoculated by skin scratch.
More stable and cheap to administer.
Occasionally causes CNS complications.
Widely used in Froncophone Africa.
Vaccination is required for travel to endemic zones (or rather, for return to
non-endemic zones!)
Lassa Fever
West Africa.
Multimammate mouse, ? vector.
Human blood is highly infectious.
Ebola
Zaire & Sudan.
No known reservoir/vector hosts.
Human blood is highly infectious - gives rise to outbreaks in hospitals and clinics,
if great care is not taken.
Marburg
(Green Monkey Disease) ? Central Africa.
Has occurred in RSA. - ? imported.
Hanta viruses
Asia, Europe, North America ? more widespread.
Rodent borne, not true arboviruses.
Droplet and rodent excreta spread.
"Hoemorrhagic Fever with Renal Syndrome" (Korean HF)
Recently: Hanta virus respiratory illness in USA.
 HERPESVIRUSES
The family of herpesviruses is very large, and its members infect most animal
species.
There are 7 herpesviruses which are known to infect humans:

• Herpes Simplex Virus type 1 and type 2

• Varicella-Zoster Virus
• Cytomegalovirus
• Epstein-Barr Virus
• Human Herpesvirus 6
• Human Herpesvirus 7

DISEASE
It is a characteristic of all herpesviruses that, following primary infection, the virus
establishes a latent infection in the host and may reactivate at any stage. Reactivation is
frequently, but not always, associated with further disease.

Structure of the virion

All herpesviruses are morphologically identical.

They have a large double stranded DNA genome. The VIRION consists of an
icosahedral nucleocapsid of about 100 nm in diameter, which is surrounded by a lipid
bilayer envelope. Between the capsid and the envelope is an amorphous layer of proteins,
termed the tegument.
 ARBOVIRUSES
and viral
HAEMORRHAGIC FEVERS

GROUP COMMON FEATURES

There is a large heterogenous group of RNA viruses (>400), which cause essentially
zoonotic infections of wild animals and are spread by insects (hence the name
arboviruses) and occasionally cause problems in man. They have the unusual ability to
multiply in the vertebrate host (usually mammals) as well as in the very different
metabolism of the insect vector.

DISTRIBUTION
They are mostly found in the warmer parts of the world (tropics) but some range into
temperate regions. Rapid transport today means every country is at some risk.

NATURE
The majority are zoonoses maintained silently in nature by their animal (vertebrate) hosts
which form their natural reservoir. In their normal hosts they have usually adapted well
and do not produce overt disease. They produce high blood titres of virus which favours
their chances of being picked up by a blood-sucking insect vector. Within that vector they
can multiply again, which favours their chances of being transmitted from the insect
vector to a new member of their definitive animal host. Crossing into non-normal hosts
may produce disease.

HUMAN INFECTION
Man usually forms an accidental, non-normal host, with sporadic - end point’ infections
which have no further transmission. However, there are some notable exceptions to this
generalisation, where human infection and disease can be spread to other humans via
human-feeding insects or even directly person to person. Human infections thus tend to
be isolated or sporadic, unless climatic conditions have favoured large-scale vector
proliferation when human (and animal) infections may become seasonal and epidemic.
Some vaccines but no specific treatments are available.
TRANSMISSION
Climate has a major role in facilitating transmission, eq. mosquito proliferation after
heavy (warm) rains. Viral replication only occurs in the insect vector in warm conditions
(>25’C). Human activity can bring man into contact with an otherwise silent cycle in
nature, eg. clearing of forests, agriculture. Creating vector promoting activities, such as
dams and irrigation, may enhance human arbovirus disease potential. Within the vector
the virus multiplies without making the insect sick and in some viruses there is evidence
there may be trans-ovarial (vertical) spread, ie. the virus can overwinter from one season
to the next inside the insect eggs. The virus is present in the insect saliva and the infection
is lifelong in the insect.

LABORATORY
Arboviruses are single-stranded enveloped RNA viruses with haemagglutinating
properties. They grow in suckling mice and/or in cell cultures. Mouse inoculation is
hazardous to human workers and must only be done in special containment facilities
(National Institute for Virology in RSA). Diagnosis is made by isolation of the
virus (usually done in a specialist laboratory) and by detection of IgM and IgG
antibodies by haemagglutination-inhibition, or ELISA.

CONTROL
Human disease control is limited to prevention by

1. control of vector, eg. mosquito control which con be very effective; and,
2. use of vaccines in a few types of infection, eg. Yellow Fever,
3. surveillance.

HUMAN DISEASE

Infection of man may produce a wide spectrum of disease - from subclinical infections to
fulminant haemorrhagic disease with death. By and large, there are 4 major clinical
patterns of disease. Any one virus characteristically tends to produce mostly one pattern,
but overlapping may occur.

Major Clinical Categories of Arboviral Disease

• No clinical illness.
• Febrile systemic illness.
• Encephalitis
• Haemorrhagic Fever.

1) No clinical illness

Severe clinical disease in man attracts attention, and many arboviruses were first
discovered (and still are today) by investigation of unexplained, possibly infectious,
disease cases. As the investigation progressed, it was commonly found that there were
milder disease cases in the vicinity of a severe disease case, aa well as subclinical cases
of infection. We now know that subclinical cases are probably more common than frank
disease in any one outbreak, eg. even in a yellow fever epidemic. New Arboviruses have
also been discovered by intensive cultural and serological investigation of possible insect
vectors (mosquitos, ticks) and possible definitive hosts (rodents, birds, small mammals,
etc.).

2. Febrile systemic illness

After inoculation there is a short incubation period of a few days, and then an abrupt
onset of disease at a time of high viraemia. There is a sudden onset of fever with chills;
headache; muscle, bone and joint pains; nausea and vomiting, and often a rash and
lymphodenopothy. These clinical features may be mild or very severe and prostrating.
The disease is typically self-limiting and short lived (few days to a week). Occasional
cases may go on to a more fulminant form (- see haemorrhagic fever section).

Examples

In South Africa.

West Nile Virus.

Asia, Central and South Africa (Transvaal).
Birds and mosquitoes. Human outbreaks after rains.
Rift Valley Fever.
East Africa, Egypt, Israel, South Africa.
Disease (with abortions) in sheep, cows, goats, spread by mosquitoes.
? occasional direct droplet spread.
Definitive host not identified.
Human outbreak follows animal outbreak after rains.
Sindbis
Africa & SE Asia (Transvaal, OFS)
Birds/mosquitoes.
Mild "Summer popular fever" epidemics in man.

Wesselsbron, Chikungunya

Other than South Africa:

Dengue.
SE Asia,, Carribean and Central America.
Mosquitoes
 Definitive host is unknown (? is none).There are seasonal epidemics in man.
4 Serological strains without cross-protection.
Previous exposure may predispose to haemorrhagic form.
(Has been imported into RSA.)

3. Encephalitis

Almost all arboviruses may infect the CNS but some characteristically produce an
encephalitis in man.
Symptoms start with fever, progressively severe headache, nausea and vomifing, neck
stiffness, paralysis, decreased consciousness, convulsions and death in severe cases.

Examples (none in South Africa)

Japanese B Encephalitis

SE Asia - seasonal.
Birds/mosquitoes - transmitted to man.
2 Human vaccines available..
(Closely related Saint Louis Encephalitis - USA, Murray Valley Encephalitis -
Australia)
Eastern Equine Encephalitis. (EEE).
Eastern USA, Central & South America.
Birds/mosquitoes.
Severe CNS disease in horses and man.
Western Equine Encephalitis. (WEE)
North & South America.
Birds/mosquitoes.
Severe disease in horses. Milder disease in man.
Tick Borne Encephalitis (TBE)
Central Europe, East Russia.
Goats/ticks.
Goats milk seems to be infectious.

4. Haemorrhagic Fever

A number of arboviruses (Toga viruses and Bunyaviruses) and some similar viruses
(Arenaviruses and Filoviruses) may cause an illness similar to the rather non-specific flu-
like illness described under (2) above, but which rapidly progresses to a severe
disseminated illness with a marked bleeding tendency and multi-organ failure.

There is usually a considerable mortality in these haemorrhagic fever infections.

Note:- Overwhelming parasitaemia/septicaemia may produce a similar clinical
picture and require to be excluded,
 eg. meningococcus, rickettsia (Tick Bite Fever), leptospirosis, hepatitis, malaria,
snake bite, tryponosomiasis.

Mild and subclinical cases also do occur, but are less easily recognised.

Examples:

In South Africa:

Crimean Congo Haemorrhagic Fever

(Eastern Europe, Central and Southern Africa)

See "What is CCHF?"

A rare disease, but the most common viral haemorrhagic fever in South Africa.
Most frequent in Kimberley area, but antibodies are widespread in cattle herds all over
South Africa.
Transmitted by ticks with striped legs (bont-poot bosluis, Hyalomma species).

Host:
cattle, ticks, ? other small mammals.
History:
visit to country in post week;
contact with animals;
tick bite;
contact with known human case (blood is highly infectious, can spread direcfly).
Illness:
short incubation (sudden onset - headache, fever, chills, body pains; diarrhoea &
vomiting;
dizzyness, confusion, abnormal behaviour,.
pharyngitis, conjunctivitis, red face and neck,
haemorrhage starts day 3 - 6 (not all cases),
multi-organ failure, that leads to death.
Treatment:
isolation and barrier nursing;
transfusions,.
ribavirin.

Outside South Africa:

YellowFever (YF)
 Africa:
Central, West East.
Central South America (Amazon basin).
Hosts: Monkey - mosquito (=silent reservoir).
Jungle (sylvan) YF
sporadic cases in forest worker/visitor.
Urban )YF
YF introduced into towns;
now get cycle: man - mosquito - man.
NB. Mosquito control is important.
Illness: as above, but jaundice is a prominent feature (liver necrosis).
Vaccine:
i) "17 D strain" live attenuated virus
Good safe long lasting protection but labile and expensive to administer.
Used in the RSA.
ii) "French Dakar Vaccine" live attenuated brain tissue-derived vaccine
Inoculated by skin scratch.
More stable and cheap to administer.
Occasionally causes CNS complications.
Widely used in Froncophone Africa.
Vaccination is required for travel to endemic zones (or rather, for return to
non-endemic zones!)
Lassa Fever
West Africa.
Multimammate mouse, ? vector.
Human blood is highly infectious.
Ebola
Zaire & Sudan.
No known reservoir/vector hosts.
Human blood is highly infectious - gives rise to outbreaks in hospitals and clinics,
if great care is not taken.
Marburg
(Green Monkey Disease) ? Central Africa.
Has occurred in RSA. - ? imported.
Hanta viruses
Asia, Europe, North America ? more widespread.
Rodent borne, not true arboviruses.
Droplet and rodent excreta spread.
"Hoemorrhagic Fever with Renal Syndrome" (Korean HF)
Recently: Hanta virus respiratory illness in USA.
 INFLUENZA
Influenza is a disease caused by a member of the Orthomyxoviridae. Many features
are common with those of the paramyxovirus infections of the respiratory tract.

CLINICAL FEATURES

Influenza is characterised by fever, myalgia, headache and pharyngitis. In addition there

may be cough and in severe cases, prostration. There is usually not coryza (runny nose)
which characterises common cold infections.

Infection may be very mild, even asymptomatic, moderate or very severe (see letter from
1918, - attached at end of page).

Source
The reservoir is acute infection in other human beings.
Spread
Is rapid via aerial droplets and fomites with inhalation into the pharynx or lower
respiratory tract.
Incubation
Is short: 1-3 days. Rapid spread leads to epidemics
Complications
Tend to occur in the young, elderly, and persons with chronic cardio-pulmonary
diseases

Consist of:
1. Pneumonia caused by influenza itself; and

2. Pneumonia caused by bacteria

- Haemophilus influenzae
- Staphylococcus aureus
- Streptococcus pneuminiae
3. Other viral superinfection, eg. Adenovirus.

Overall death rates increase in times of influenza epidemics.

LABORATORY DIAGNOSIS

A. Viral Isolation:

Respiratory secretions:
- direct aspirate
- gargle
- nasal washings

a) Rapid examination of cells by immunofluorescence.

b) Inoculation of cell cultures (or eggs).

B. Serology
serum antibodies by haemagglutination inhibition

INFLUENZA VIRUSES

Diagramatic representation of the morphology of an influenza virion.

The virion is generally rounded but may be long and filamentous.

A single-stranded RNA genome is closely associated with a helical nucleoprotein (NP),
and is present in eight separate segments of ribonucleoprotein (RNP), each of which
has to be present for successful replication. The segmented genome is enclosed within an
outer lipoprotein envelope. An antigenic protein called the matrix protein (MP 1) lines
the inside of the envelope and and is chemically bound to the RNP. The envelope carries
two types of protruding spikes. One is a box-shaped protein, called the neuraminidase
(NA), of which there are nine major antigenic types, and which has enzymic properties as
the name implies.
 The other type of envelope spike is
a trimeric protein called the haemagglutinin (HA)
(illustrated on the right)
of which there are 13 major antigenic types. The haemagglutinin functions during
attachment of the virus particle to the cell membrane, and can combine with specific
receptors on a variety of cells including red blood cells.
The lipoprotein envelope makes the virion rather labile - susceptible to heat, drying,
detergents and solvents.

REPLICATION

The Life Cycle of Influenza Virus

Receptor-bound viruses are taken into the cell by

endocytosis. In the low pH environment of the
endosome, RNP is released from MP1, and the viral
lipoprotein envelope fuses with the lipid-bilayer of
the vesicle, releasing viral RNP into the cell cytoplasm, from where it is transported into
the nucleus. New viral proteins are translated from transcribed messenger RNA (mRNA).
New viral RNA is encased in the capsid protein, and together with new matrix protein is
then transported to sites at the cell surface where envelope haemagglutinin and
neuraminadase components have been incorporated into the cell membrane. Progeny
virions are formed and released by budding.
The cell does not die (at least not initially).

Flu is one of a rare few viruses that has its genome in separate segments (eight). - This
increases the potential for recombinants to form (by interchange of gene segments if two
different viruses infect the same cell), and may contribute to the rapid development of
new flu strains in nature - can also be duplicated in the laboratory (used for making
vaccine strains). Avian and human strains recombining in pigs in the Far East may
permit virulent human strains to evolve.

CLASSIFICATION of virus STRAINS

Is done on the basis of antigenicity of NP and MP into three main groups:

Influenza A -HA undergoes minor and occasional major changes - very important.
- NA some variation.
Influenza B) Undergoes relatively slow change in HA with time. Known only in man.
Influenza C) Uncommon strain, known only in man.

NOMENCLATURE
Influenza strains are named in the following way:
A SINGAPORE 6 86 (H1N1)
TYPE TOWN NUMBER YEAR MAJOR
of influenza where first isolated of isolates of isolation TYPE of HA and NA

EPIDEMIOLOGY

Influenza A virus is essentially an avian virus that has "recently" crossed into mammals.
Birds have the greatest number and range of influenza strains. Avian haemagglutinins
sometimes appear in pig human and horse influenza strains.
Every now and then (10 - 15 years) a major new pandemic strain appears in man, with a
totally new HA and sometimes a new NA as well (antigenic shift). This variant causes a
major epidemic around the world (pandemic).
Over the subsequent years this strain undergoes minor changes (antigenic drift) every
two to three years, probably driven by selective antibody pressure in the populations of
humans infected.
See chart below indicating main pandemic strains in previous years.

Influenza A Evolution
1874 --- (H3N8)
1890 --- (H2N2) .........................Pandemic
1902 --- (H3N2)
1918 --- (H1N1)..........................Pandemic
1933 --- (H1N1)..........................First strains isolated
1947 --- (H1N1)..........................Variation detected
1957 --- (H2N2).........................."Asian" Flu pandemic
1968 --- (H3N2).........................."Hong Kong" Flu pandemic
1976 --- (H1N1).........................."Swine" Flu, non-epidemic
1977 --- (H1N1) + (H3N2)........."Russian" Flu epidemic

This constant antigenic change down the years means that new vaccines have to be
made on a regular basis.

New influenza strains spread rapidly in children in schools and créches and in places
where people crowd together. Influenza epidemics may cause economically significant
absenteeism.

TREATMENT

Antibiotics are often prescribed - have no effect on virus but may prevent or cure
bacterial superinfection. The drug Amantadine may prevent influenza if taken
continuously by high-risk persons at the time of an epidemic, but is not used widely.

PREVENTION

Vaccines at best give about 70% protection.

They may sometimes not be effective against the most recently evolved strains because
the rate of evolution outpaces the rate at which new vaccines can be manufactured.

Types of Vaccine

Killed Whole Virus

Rather pyrogenic, not used today.
Live Virus
Attenuated strains were widely used in Russia but not elsewhere.

Virus Subunit
HA extracted from recombinant virus forms the basis of today's vaccines.
For example, the WHO Recommendation for Influenza Vaccine, 1995-1996, contains
two A strains and one B strain:-

A / Singapore / 6 / 86 (H1N1)
A / Johannesburg / 33 / 94 (H3N2)
B / Beijing / 84 / 93

Synthetic
Much research is being done to try and find a neutralising epitope that is more stable, and
can therefore be used for a universal vaccine.

PROSPECTS

Because another devastating pandemic strain (cf 1918 pandemic) may appear at any time,
the World Health Organisation (WHO) maintains worldwide surveillance of flu strains
and makes predictions of suitable strains for vaccine production.

MUMPS
Mumps virus causes a febrile illness and has a tropism for glandular tissue. It is also
relatively neuroinvasive.

Mumps virus is spread by the respiratory route, and has a relatively long incubation of
about 21 days. It causes a febrile illness and inflammation of the salivary glands,
classically the parotid and submaxillary glands. The swelling of the glands may be
asynchronous, and lasts about 1 week.

Aseptic meningitis is a fairly common complication of mumps, and by corollary, mumps

is a fairly common cause of aseptic meningitis (up to 1/3 of cases in Cape Town). In
about half of the mumps meningitis cases, parotitis will not be apparent.

Mumps meningoencephalitis is rarer but a more serious development.

Orchitis can occur, more often after puberty, unilateral or bilateral, but is rarely followed
by infertility. Other glandular tissue is very occasionally involved viz. pancreatitis,
oophoritis or thyroiditis.
Diagnosis can be confirmed by positive IgM antibodies. In cases of meningitis the virus
is readily isolated from cerebro-spinal fluid.

Mumps is a paramyxovirus, the virus family with the tubular spaghetti-like

nucleocapsid. Measles is also a paramyxomvirus. The other human paramyxoviruses
cause respiratory infections - they are the parainfluenza viruses and respiratory syncitial
virus.

Mumps is considered as a common old garden infection that you may have had or seen
someone with. It causes swelling of the salivary glands, especially the parotids. The
swelling is usually bilateral, and can be uneven, ie. one side starts before the other. The
child will already have been a bit feverish and unwell for a few days. Now mumps virus
fancies glandular tissue, and the next most common gland that it affects, after the salivary
glands, is the testis; so a common and unpleasant complication in boys after puberty is
orchitis. Contrary to old wives tales, it is rare to become infertile from mumps orchitis.
The ovaries can also sometimes be affected causing oophoritis; and the pancreas causing
pancreatitis.

Apart from glands, mumps virus crosses the BBB (blood brain barrier) rather easily, and
if you did a lumbar puncture on every child with mumps you would find that 10% of
them had meningitis, even if they didn't have overt symptoms. Mumps meningitis is
common - it causes about 1/3 of viral meningitis in Cape Town, but it has a good
outlook. Sometimes the virus goes further and actually invades the brain, ie
meningoencephalitis. This is a bit more serious, and the most common long-term
consequence is unilateral deafness.

For suspected mumps meningitis it is quite easy to grow the virus from cerebro-spinal
fluid.
 NEUROLOGICAL DISEASES CAUSED BY VIRUSES

Neurological disease is one of the most serious complications of virus infection.

Clinically, viral neurological disease can be divided into
acute diseases and chronic syndromes.
The pathology may be due either to viral multiplication in the cells of the brain, or due
to the (misdirected) immune response of the host - post infectious encephalo-myelitis.
Where virus replicates in the brain, virus can usually be isolated from brain tissue or from
cerebrospinal fluid. This is not the case with the post infectious syndromes.
 Acute neurological syndromes

Virus may reach the brain either by the blood stream or by spread along peripheral
nerves. Asymptomatic infection of the brain is common.

There are four main syndromes:

• 1. Encephalitis
• 2. Flaccid paralysis
• 3. Aseptic meningitis
• 4. Post infectious encephalo-myelitis

Acute viral encephalitis

Viral replication occurs in the brain tissue itself, causing destructive lesions in the grey
matter. The main symptoms include: fever, drowsiness, confusion, convulsions and focal
neurological signs. Morbidity and mortality is very high.
Viruses that cause encephalitis include
Herpes Simplex, Rabies and some of the Arboviruses.

The Arboviruses include a miscellaneous group of enveloped, RNA viruses, that are
transmitted from one vertebral host to another via blood sucking arthropods. The main
reservoirs are wild birds and small mammals. Man may be infected if bitten by the insect
vector. Arboviruses cause two types of disease:

• 1. Encephalitis, and
• 2. Fever (often with haemorrhagic manifestations)

Arboviral encephalitis occurs in most parts of the world; but different agents are
responsible for disease in different areas. In South Africa, there are no endemic
arboviruses that specifically cause encephalitis. Rarely, however, encephalitis may occur
as part of the clinical course of infection with: West Nile virus, Rift Valley Fever virus,
Sinbis virus, Crimean-Congo haemorrhagic fever or Chickengunya virus. These
viruses are endemic in livestock herds in certain parts of the country and farm workers or
vets may occasionally be infected.

Flaccid Paralysis

Due to direct infection of motor neurones. Patients present with fever and flaccid
paralysis of a group of muscles. The lower limbs tend to be involved more commonly
than the upper limbs. Signs of meningitis such as headache and neck stiffness are
frequent accompanying features. The most common aetiological agents include the
polioviruses 1-3, but with the reduction in prevalence of wild type polio, a number of
non-polio enteroviruses have also been implicated as rare causitive agents.

Aseptic meningitis

Infection of the meninges. A relatively mild disease with a good prognosis. Patients
present with fever, headache, neck stiffness and photophobia. Common viral agents
include: enteroviruses, mumps virus and lymphocytic choriomeningitis virus.

Post infectious encephalomyelitis

This uncommon complication may develop in the convalescent phase of a number of

common virus infections, including: measles, mumps, rubella and primary varicella-
zoster virus infection, as well as following the administration of certain vaccines, such
as: vaccinia virus and the older neurotissue rabies vaccines. Wide spread demyelinating
lesions develop in the brain and spinal cord, associated with lymphocytic infiltration
and perivascular cuffing of adjacent blood vessels. Virus cannot be isolated from brain
tissue or CSF. The aetiology is somewhat obscure, but it is believed to be an auto-
immune phenomenon, triggered by exposure to foreign antigens which are closely
related to host proteins normally expressed in brain tissue (molecular mimicry).

Gillain Barre syndrome

Poly-neuritis which develops a few days after the acute phase of a viral infection. The
disease is due to demyelination of peripheral nerves. Patients present with an ascending
paralysis, associated with paraesthesia. Like post infectious encephalomyelitis, it is
believed to be an immunological phenomenon. Patients usually recover spontaneously
over a few weeks or months. There is no specific treatment.

Chronic Neurological Conditions:

A number of viruses and virus-like agents can cause chronic neurological disaese.
Characteristically, these conditions have a long incubation period followed by slow
development of symptoms and a progressive, uniformly fatal course.
 • Subacute-sclerosing panencephalitis (SSPE)
• Progressive multifocal leuco-encephalopathy (PML)
• Retrovirus disease
• Spongiform encephalopathies

Subacute Sclerosing Pan Encephalitis

This is a rare, slowly progressive degenerative disease of the brain which develops six to
eight years following a primary uncomplicated infection with measles virus. It is due to
persistent measles virus infection in the CNS.

Clinical Features
Patients usually present with personality and behavioural changes followed by
progressive intellectual impairment, convulsions, myoclonic movements, coma and
death.

Pathogenesis
It is thought that the virus gains entry into the CNS during the viraemia that occurs at the
time of the primary infection. The pathology is due to infection of the brain with a
measles virus mutant that lacks an essential structural gene and cannot therefore undergo
complete cycles of replication. This has two effects:

1.) It enables the virus to establish a persistent infection in the brain - because there is
limited expression of viral antigens on the surface of infected neurones, an effective
immune response cannot be mounted against the infected cells.

2. ) The disease proceeds very slowly - because, as no infectious virus particles are
released, spread of infection to new cells only occurs through fusion with adjacent
infected cells.

Epidemiology
Occurs in about 1 per million cases of measles. The incidence has declined sharply since
the introduction of vaccination against measles.

Laboratory diagnosis
Affected individuals have high titres of measles specific antibodies in their serum and
CSF.
Measles virus antigens can be demonstrated in brain tissue

Progressive multifocal leuco-encephalopathy

 There are two members of the Papovavirus family which are known to infect man,
namely JC virus, which replicates in neural tissue and BK virus which has been isolated
from the urinary tract. Neither of them cause any pathology in healthy people.
JC virus - is associated with progressive multifocal leuco-encephalopathy

Virology

The Papovavirus family contains two main groups:-

Papilloma viruses (which cause warts), See electronmicrographs

Polyoma viruses,

these are slightly smaller than papilloma viruses, and in animals are sometimes
associated with tumours.
The two known human polyoma viruses are:
JC - Progressive multifocal leuco-encephalopathy
BK - isolated from urine of immunosuppressed patients

dsDNA genome

small icosahedral particles, 42-45 nm

grows in human foetal glial cell cultures (JC) Progressive multifocal leuco-
encephalopathy is a progressive neurological disorder caused by reactivation of JC
virus in the brain. Infection is common, but neurological disease is rare; it only occurs
in immunosuppressed patients.

Clinical Features
Patients may present with a variety of neurological signs, including: hemiparesis,
dementia, dysphagia, muscular inco-ordination or impaired vision. The condition is
progressive and invariably fatal.

Pathology
Multiple foci of demyelination are found throughout the cerebral hemispheres and
cerebellum. The virus infects oligodendrocytes, which have a bizzare histological
appearance.

Epidemiology
Infection with JC virus is common, but invariably asymptomatic: Seroprevalence surveys
have shown that about 50-60% of adults have antibodies.

Spongiform Encephalopathies
(Prion diseases)
A number of transmissable neurological syndromes, caused by unconventional virus-like
agents, have been identified in man and other animals. These diseases are characterized
by the following features:

1. Confined to the CNS

2. Long incubation period
3. Progressive, uniformly fatal course
4. Typical brain histology: reactive gliosis, vacuolation of neurones, deposition of
amyloid protein in the brain and the absence of an inflammatory response.
5. Infection is believed to be transmitted by means of a novel infectious protein,
termed prion or PrP.

Diseases falling into this category include:

Scrapie - sheep, goats
Bovine spongiform encephalopathy - cattle
Transmissable Mink encephalopathy - mink
Creutzfeldt-Jakob disease - man, sporadic and familial
Gerstmann-Streussler disease - man, familial
Fatal Familial Insomnia - man, familial
Kuru - man, Fore people, New Guinea

Properties of Prions:

• 1. Infectivity is associated with a novel infectious protein, termed prion (PrP)

• 2. Infectious particles are very small - about 20-30 nm in size.
• 3. No nucleic acid has yet been found to form part of the agent
• 4. Prions are extremely resistent to inactivation by: ultra-violet light,
formaldehyde and heat.
• 5. Prions can be transmitted by intra-cerebral or sub-cutaneous inoculation of
material from infected brain.

Pathogenesis of prion diseases

The pathogenesis of prion diseases is still a great mystery. The pathology appears to be
both infectious as well as genetic.

Recent evidence has demonstrated that the prion protein has the same primary amino
acid sequence as a host protein which is normally present at low levels in healthy brain
tissue. The cellular protein has been termed PrPc. Although the amino acid sequence is
the same as the cellular one, the prion protein, has an abnormal conformation, due to
different post translational modification. Exposure to the abnormal form triggers a
conversion of the cellular isoform to the infectious form. This is followed by progressive
accumulation of the abnormal form (prion protein) in the brain, which is deposited as
amyloid.

Scrapie
Scrapie is a neurological disease occurring in sheep and goats. It was first identified in
Britain, but it is now known to occur world wide. It is the prototype disease of the
spongiform encephalopathies; the scrapie prion is termed PrPsc. The disease is clearly
infectious as it can be transmitted to sheep, mice and hamsters by intra-cerebral or sub
cutaneous inoculation of material from the brains of infected sheep. Susceptibility to
infection appears to be genetically determined.

Clinical features
The disease is transmitted by contact in flocks of sheep and also vertically, from ewes to
lambs. In natural infection, the incubation period is 2 to 5 years. Following intra-
cerebral inoculation, however, the incubation is much shorter, only 3 to 24 months.

Affected sheep develop ataxia, tremor and muscular in co-ordination; the symptoms
progress to paralysis and death.

Bovine Spongiform encephalopathy

Similar disease to scrapie which occurs in cattle. It is believed that scrapie was
introduced into cattle herds fed on feeds containing scrapie-infected sheep offal. The
condition is now epidemic in cattle herds in England and Wales ("Mad Cow Disease").
There is concern that the infection may be transmitted to humans, and a report of an
increased number of cases of CJD in young people linked to the consumption of beef
burgers sparked a world wide furore.

Transmissable Mink encephalopathy

Scrapie has been introduced into minks, bred in captivity through feeds containing sheep
brains.

Creutzfeldt-Jakob disease (CJD)

In 1920, Creutzfeldt described a progressive dementing illness in a 22 year old woman.

The following year, Jakob described four older patients with a clinically similar
presentation and course. Since then, numerous cases of CJD have been described. CJD
occurs world wide. It is very rare, with an incidence of about 1 case per million
population. While most cases are sporadic, 5-10% are familial. In the familial form, CJD
is inherited as an autosomal dominant condition.

Clinical features
The onset of the disease typically occurs between the ages of 50 and 65 years. There are
two main modes of presentation:

• 1. Chronic dementing illness

• 2. Progressive cerebellar dysfunction

The condition is relentlessly progressive and patients usually die within a year of
presentation.

Transmission:
The natural route of infection is not known, but CJD has been accidently transmitted by:
1. Corneal grafts - where the corneas were harvested from cadavers that died of CJD.
2. Growth hormone preparations, derived from human pituitary glands.
3. In two patients who received grafts of dura mater, prepared from cadavers, and
4. through electrodes used for electro-encephalography which had previously been used
in a patient with CJD.

The prion detected in the brains of patients with CJD is termed PrPCJD. The disease has
also been experimentally transmitted to chimpanzees and other primates. Following
intracerebral inoculation, the incubation period is 11-14 months. The disease can also be
transmitted peripherally (IV, intra peritoneal or intramuscular routes). But transmission is
much less efficient and the incubation period is much longer (many years).

Decontamination procedures:
Prions are extremely resistent to inactivation by ultra-violet light, forrmaldehyde and
heat.

Recommended methods of decontaminating infected material include:

1. Autoclaving for 4.5 hours (121oC, 15 p.s.i)
2. 1N NaOH followed by autoclaving for 1.5 hours

Gerstmann-Streussler Syndrome

Rare familial neurodegenerative disease which usually manifests in the third to the
seventh decade of life. The condition is both infectious and genetic: the predisposition is
inherited as an autosomal dominant condition; while, in addition, the disease can be
transmitted to non human primates by intra cerebral inoculation with brain tissue from
cases of GSS.
Recent genetic studies have revealed that affected individuals have a point mutation in
their PrPc gene. The altered amino-acid sequence of the PrPC protein may make it more
susceptible to transformation to the abnormal conformation.

Fatal Familial Insomnia

Another rare familial disorder. Predisposed individuals also have a mutation in their PrPc
gene.

Kuru

This is a transmissable prion disease found only in the Fore people of New Guinea. It
first appeared about 60 years ago and the incidence increased until the late 1950's. The
disease is now known to have been transmitted through ritual cannibalism: until the late
1950's it was the practice of women and children to eat the brains and viscera of dead
relatives, including those who had themselves died of Kuru. Cannibalism stopped in 1957
and the incidence of Kuru has declined sharply since then.

Clinical features:
The incubation period varied from 4-20 years.
Patients presented with progressive cerebellar dysfunction. Death usually occurred within
a year of initial presentation.

Transmission studies:
Intra-cerebral inoculation of brain tissue from Kuru victims into non human primates
leads to the development of symptoms within two years.

Retrovirus Disease

Lentivirinae:
Lentiviruses characteristically establish a persistent infection in the host and cause
chronic wasting disorders which are uniformly fatal. Members of this family that
typically cause CNS pathology include:

Virus Host Clinical Features

Visna-Maedi virus sheep paralysis, wasting, ataxia
Caprine arthritis encephalitis virus (CAEV) goats paralysis, wasting, ataxia
HIV 1 and 2 humans dementia
Pathology
Inflammatory cell infiltration, perivascular cuffing of blood vessels, demyelination,
necrosis and reactive gliosis.

Pathogenesis
The pathogenesis of the neuropathology is not very clear. Virus is probably introduced
into the CNS by infected monocytes which cross the blood brain barrier. Differentiation
of the infected monocytes into microglial cells is thought to trigger viral replication.
None of these viruses appear to infect neuro-ectodermal cells directly, and damage to
brain tissue is therefore thought to occur indirectly by cytokines released during
inflammation. It is also thought that certain viral proteins, such as the gp160 of HIV, may
be directly toxic to neurones.

In addition to the above ( where the neurological damage arises directly as a result of
HIV infection) CNS pathology in patients with AIDS may also be caused by
opportunistic infectious agents such as: JC virus (PML), HCMV, VZV as well as a
number of bacterial and fungal diseases

Oncovirinae

HTLV 1 - Tropical spastic paraparesis

HUMAN PAPILLOMAVIRUSES
(WART VIRUSES)
FAMILY: PAPOVAVIRIDAE
GENUS: PAPILLOMAVIRUS

CLINICAL FEATURES AND VIROLOGY

Papillomaviruses occur in many animals as well as man but have narrow host specificity
and tissue specificity. Human papillomaviruses do not infect any animal hosts.

In humans, HPV is one of the most common viral infections. Spread is by contact, eg.
hand to hand; sexual transmission in the case of genital infections; infection during
birth in the case of juvenile laryngeal papillomas.

There are more than 80 different types of HPV. Typing is based on the DNA sequence of
the virus. Specific lesions are associated with specific HPV types. There are three major
groups of HPVs.
1. Cutaneous warts
2. Epidermodysplasia verruciformis
3. Mucosal HPV infections

VIRUS STRUCTURE

The papillomaviruses are small non-enveloped icosahedral viruses (55nm) and have a
circular double stranded DNA genome of about 8kb.

LIFE CYCLE OF HUMAN PAPILLOMAVIRUS

HPVs infect the germinal epithelial cells through damaged epithelium. The viral DNA
replicates in the lower layers of squamous epithelium. However, expression of the late
genes coding for the viral capsid proteins is linked to the differentiation of the epithelial
cells and viral particle are only detectable in the upper layers of the epithelium. Therefore
if the cells fail to differentiate as observed in malignancy, there is a non-productive
infection. This is the reason why it was only when molecular techniques were developed
for the detection of HPV DNA that it became possible to link HPV and cervical
carcinoma.

Cutaneous warts
(HPV types 1-4, 7, 10, 26-28)

These viruses cause warts (benign tumours) of keratinised squamous epithelium, ie. skin.
They are very common and the peak incidence is in children between the ages of 10 and
14 years of age. The cutaneous group of viruses has traditionally been classified
according to clinical appearance and location. There are four clinical types:

• verruca vulgaris or common wart

• deep hyperkeratotic palmoplantar wart or myrmecia
• superficial mosaic type palmoplantar wart
• verruca plana or flat wart

In most people skin warts regress spontaneously.

Epidermodysplasia verruciformis
(HPV types 5, 8, 9, 12, 14, 15, 17, 19-25)

This is a rare life-long disease associated with specific HPV types. There appears to be a
genetic component to the disease as large proportions of people with the disease are
descendants of consanguineous marriages. It is probably an autosomal recessive based
inheritance. The infected people develop widespread warts with about 30% possibility of
the lesions progressing to malignancy. Malignancies are most likely to develop in sun-
exposed and traumatised area.

Mucosal HPV lesions

These HPV viruses infect the genital tract, oral cavity, conjunctiva and respiratory tract.
The majority of infections are latent with no visible lesions.
Table 1 lists the major HPV types associated with mucosal HPVs.

HPVs are also the most common sexually transmitted viruses. It is estimated that there
are 450 000 new cases of cervical cancer worldwide, 10 million cases of high grade
dysplasia, 30 million cases of low grade dysplasia and 300 million cases of cervical HPV
infection without cytological abnormalities (IARC, 1999). About 200 000 women will
die of cervical cancer each year.

Condylomata acuminata (genital warts): This is the most common benign genital tumour
found in both sexes. These tumours hardly ever progress to malignancy. In the USA and
UK, 1-3% of sexually active people have genital warts.

Laryngeal papillomas: This is a rare condition found in both adults and children. The
juvenile laryngeal papillomas are a result of transmission from the mother to the infant
during birth. The same HPV types are found in laryngeal papillomas and genital warts.
Although this is a benign tumour they have to be removed on a regular basis to prevent
blockage of the respiratory tract.

Table 1. Major clinical association of genital tract and other mucosal HPVs (from Fields
et al., 1996)

CLINICAL ASSOCIATION HPV TYPES

GENITAL TRACT
Subclinical infection All types
Exophytic condyloma - any site HPV 6 , 11
Flat condyloma - especially cervix HPV 6, 11,16, 18, 31, others
Bowenoid papulosis HPV 16
Giant condyloma HPV 6, 11
Cervical cancer
• Strong association
HPV16, 18, 31, 45
• Moderate association
HPV 33, 35, 39, 51, 52, 56, 58, 59, 68
• Weak or no association
HPV 6, 11, 26, 42, 43, 44, 53, 54, 55, 62
Vulvar cancer HPV 16
Penile cancer HPV 16

RESPIRATORY PAPILLOMAS HPV 6, 11

CONJUNCTIVAL PAPILLOMAS HPV 6, 11

ORAL CAVITY
Focal epithelial hyperplasia HPV 13, 32
Infection with genital tract HPVs HPV 6, 11, 16
Lesions on Lip HPV 2
 Cervical cancer and other cancers:

A significant proportion of cancers is associated with HPV infection: 11% of all cancers
in women and 2% of all cancers in men. South Africa has one the highest incidences of
carcinoma of the cervix in the world. This is particularly true in our disadvantaged
communities. The malignant and premalignant lesions (cervical intraepithelial neoplasia)
can be detected by routine cervical screening (PAP smears). If detected early, treatment is
very successful. The high incidence of the disease in South Africa is mainly due to poor
cervical screening programmes.

It is now accepted that specific types of HPV play a causal role in cervical cancer. There
is epidemiological evidence that cervical cancer is associated with a sexually transmitted
agent. This includes strong risk factors such as early age of first intercourse, multiple
sexual partners, high parity etc. The disease is virtually unknown in nuns and virgins and
is relatively common in commercial sex workers. Over the years a number of sexually
transmitted agents have been investigated as possible aetiological agents for cervical
cancer. The strongest association has been found between specific HPV types and
cervical cancer. HPV types 16 ,18, 31 and 35 are most commonly found in cervical
cancer biopsies with HPV 16 being found in half of the cervical cancer cases worldwide.
They are also found in penile and anal carcinomas. Recent studies also support the
hypothesis that cervical intraepithelial neoplasia is caused by HPV. HPV DNA can be
detected in over 95% of cervical cancer and intraepitheial neoplasia biopsies.

MOLECULAR BASIS OF HPV INDUCED GENITAL CANCERS

Experimental evidence supports the epidemiological findings that specific HPV types do
play a role in progression to malignancy.

HPV types associated with malignancy (HPV 16 and 18) have different characteristics
compared with those types associated with benign lesions (HPV 6 and 11). One of the
important steps in progression to cancer is the disruption of the activity of two host
tumour suppressor proteins; namely p53 and the retinoblastoma protein (pRb). Tumour
suppressor gene products have the normal task of halting cell growth. The HPV E6 gene
product binds p53 and the HPV E7 gene product binds pRb. However the E6/E7 proteins
of HPV 16 and HPV 18 have a much higher affinity for the tumour suppressor gene
products than the E6/E7 proteins of HPV 6 and 11. This difference helps explain the
difference in the transforming abilities of the virus associated with benign and malignant
lesions. If the function of p53 is disrupted then the cell cycle does not pause to allow
repair of damaged DNA. This results in an accumulation of mutations in the cells and it is
postulated that eventually one of the proto-oncogenes will mutate. Proto-oncogenes are
genes involved in normal cell growth and division. When mutated they promote
uncontrolled cell growth and are known as oncogenes.

In the "normal" life cycle of the virus the production of E6 and E7 are controlled by the
E2 gene product. However in cancers the E2 gene is disrupted during integration of the
viral genome into the host cell. This results in uncontrolled production of E6 and E7 for
interaction with the tumour suppressor gene products. Once again integration is mainly
observed with the HPV types associated with malignancy ie HPV 16 and 18.

It is important to realise that very few women infected with HPV will progress to cervical
carcinoma. In a study done at UCT, 13% of women with normal cervical cytology were
found to be infected with HPV. However, the incidence of cervical carcinoma in the
Black population is 38 per 100 000. Cancer is a multistep process and HPV is only one of
the steps in the progression to cancer. Several co-factors have been identified including
immunosuppression, smoking and the presence of other sexually transmitted diseases.

DIAGNOSIS OF HPV INFECTION

- No serological diagnosis
- No convenient cell culture
- Cytology
- Histology
- Colposcopy in the case of genital HPV infections.
- Electron microscopy. Only applicable in benign lesions as in malignant lesions the viral
DNA is integrated and viral particles are not produced.
- Immunocytochemistry can detect major capsid protein but are generally group specific
not type specific
- DNA detection techniques. This is the only way to type HPVs. Typing is considered to
be important, as specific HPV types are associated with malignancy. Polymerase chain
reaction can be used to detect HPV DNA. At present the only commercial test available is
the "Hybrid CaptureTM" test which will differentiate between oncogenic and non-
oncogenic HPV infections.

PREVENTION

In the future there will be vaccines to prevent HPV infection (prophylactic vaccination).
At the moment only Phase One trails have taken place so it will not be within the next ten
years. There is also research taking place to develop therapeutic vaccines to treat people
infected with HPVs.
 RUBELLA and
PARVOVIRUS B19
RUBELLA and PARVOVIRUS B19 are world-wide childhood illnesses caused by
unrelated viruses:

Rubella virus and parvovirus B19 cause similar, mild rash diseases, and arthritis is a
common complication of both. Both may endanger the foetus if the mother is infected in
pregnancy, but in very different ways.

Parvovirus B19 has other unique complications in children and adults because of its
tropism (predilection or affinity) for immature red cells.

For both of these viral infections, man is the only natural host and reservoir, so that the
viruses must keep circulating in human populations, and tend to occur in small
epidemics. However, because of long-lasting immunity, these infections usually occur
only once in one person's lifetime .
Note that the person is infectious for about a week before the onset of symptoms, and
then for a few days from the onset of symptoms.

RUBELLA

Rubella virus is shed in oropharyngeal secretions and is spread by the respiratory route.
After a 2 - 3 week incubation, a rash of pink macules appears on the face and behind the
ears, and spreads downwards to the trunk and limbs. There is associated low-grade fever
and lymphadenopathy, specifically of the posterior cervical and occipital nodes. These
signs resolve in a few days. Arthralgia / arthritis, usually of the fingers and knees, can
occur, especially in adult women.

It is relatively common for rubella infection to occur without a rash, so called

"subclinical" infection.

Post-infectious encephalitis or thrombocytopaenia are very rare complications.

Congenital rubella syndrome

Primary rubella infection of the mother in the first 12 weeks of pregnancy is very likely
to infect the foetus, with a high risk (80%) of congenital abnormalities. As in other
congenital infections, the baby at birth may be of low weight, and have a
thrombocytopaenic rash and hepatomegaly with jaundice.

Specific features of congenital rubella syndrome are: cataracts, micropthalmia, heart

defects, sensorineural deafness, mental retardation. Infection between about the 13th and
the 17th week of pregnancy may result in deafness alone, but infection beyond the 17th
week is no longer a hazard.

A confirmed laboratory diagnosis of rubella in a pregnant woman is thus very important,

as termination should be offered for 1st trimester infections.
Rubella IgM by ELISA becomes detectable a few days after the rash appears. The
presence of IgM antibodies indicates an acute infection, and will be detectable for
approximately 8 weeks. (Rubella IgG will appear almost as early as the IgM, but then
persists for life).
Suspected congenital rubella in a neonate is also diagnosed by IgM in the baby's blood,
or by culture of the virus from urine.
Note that maternal IgG class antibodies can cross the placenta, but not IgM, and so the
presence of IgM in the baby indicates production of its own antibodies in response to its
own infection. Congenitally infected babies may shed virus for many months, and staff
and family contacts should be aware that the baby is infectious for this time.

PREVENTION:

Rubella vaccine is not given routinely in childhood in South Africa. The vaccine is a live
attenuated strain called RA27/3. In some countries it is given as the combined MMR
(measles, mumps, rubella - all live, attenuated) vaccine to all children at 15 months of
age, with a second dose at school entry. Some countries have tried a policy of rubella
vaccine alone given to girls at high school entry age (+/-13 years) specifically to try to
prevent congenital rubella. In practice, good herd immunity of both boys and girls is
necessary to prevent circulation of the virus and protect vulnerable pregnant women.
Women should be screened for immunity to rubella (indicated by IgG antibodies) prior
to pregnancy so that they can be vaccinated if necessary.
(Note that live virus vaccines should not be given during pregnancy, but inadvertent
rubella vaccination during pregnancy is not grounds for termination as there have
been no cases of congenital abnormalities associated with the vaccine.)
Often women are only screened during their first pregnancy, but they can at least be
vaccinated prior to the next pregnancy if they are non-immune.

PARVOVIRUS B19

This virus was only discovered in the 1970's, although the disease it causes, erythema
infectiosum, has long been described. It is also sometimes called "fifth disease" from an
historical enumeration of the rash diseases of childhood, or "slapped cheek disease"
because of the facial rash. Its name "B19" refers to the batch number of a serum in the
bloodbank in which the virus was discovered.

Parvo B19 is also spread by the respiratory route, and the rash appears after about 17
days. It's distinctive feature is the red cheeks, with circumoral pallor. On the rest of the
body it is a lacy, pink macular rash that fades quickly, but may reappear after a warm
bath. Arthralgia or frank arthritis, in fingers and knees most commonly, is more frequent
and troublesome than in rubella. The arthritis can in some cases persist for months, and
can even imitate juvenile rheumatoid arthritis.

Parvo B19's site of replication is red cell precursors in the bone marrow. It utilises as its
receptor the P antigen (a red cell surface glycoprotein) which determines the P blood
group phenotype.
The infection will cause a temporary shut-down in red cell production until the virus is
eliminated by the immune system, usually within 10 days. In a normal child this causes
an insignificant drop in haemoglobin of around 1g/dl. However, people with hereditary
red cell disorders (eg sickle cell anaemia, hereditary spherocytosis, thalassaemia) have
either red cell under-production or rapid red cell destruction by haemolysis. In this
context, the brief cessation of red cell supply caused by parvo B19 infection will
precipitate an "aplastic crisis" ie. severe anaemia. These patients present with extreme
pallor, lethargy and sometimes in cardiac failure, and require blood transfusion. A parvo
B19 vaccine will soon be available and will benefit this vulnerable group.

In persons with AIDS, or other immunodeficiency states eg. children with leukaemia on
chemotherapy, inability to clear the virus can cause chronic anaemia.
Normal immunoglobulin preparations for intra-muscular injection contain parvo B19
antibodies and will usually successfully eliminate the infection in immuno-compromised
patients.

Maternal infection with parvo B19 in pregnancy can cause foetal infection, and foetal
anaemia. In the worst cases (rare, usually second trimester infection) this may manifest
as foetal hydrops. The mechanism here is severe foetal anaemia causing cardiac failure
with oedema. This may end in intra-uterine death, but it has been possible to treat
severely affected foetuses by intra-uterine transfusion, given the right technology. The
milder cases tend to resolve spontaneously.

Note that parvovirus B19, unlike rubella, is not teratogenic, and outcome of infections in
pregnancy is usually good.

Because the rash and mild illness cause by parvo B19 is very similar to rubella, it is
essential in pregnant women to distinguish between the two virus infections by laboratory
diagnosis.
Laboratory diagnosis of acute parvo B19 is also based on the presence of IgM antibodies.
The virus cannot be cultivated in routine cell culture lines, but direct detection of the viral
DNA may be achieved by PCR.

POX VIRUSES
INTRODUCTION

A range of pox viruses cause febrile illnesses in man and animals with a prominent
vesicular rash. The most prominent of them was smallpox virus (variola) which caused
a severe disease in man but which has now been eliminated by intensive international
vaccination.
Current interest in poxviruses centers around their possible use as vaccine vectors.
THE VIRUS

Smallpox belongs to the Orthopox virus genus. Poxviruses are very large, brick-shaped
viruses about 300 x 200 nm (the size of small bacteria). They have a complex internal
structure - a large double-stranded DNA genome (about 200 kbp in size) is enclosed
within a "core" that is flanked by 2 "lateral bodies". The surface of the virus particle is
covered with filamentous protein components, so that the particles have the appearance
of a "ball of knitting wool". The entire particle is enclosed in an envelope derived from
the host cell membranes. Most poxviruses are host-species specific, but vaccinia is a
remarkable exception.
True pox viruses are antigenically rather similar, so that infection by one elicits immune
protection against the others.
Laboratory diagnosis of pox viruses may be undertaken by electron microscopy of
negatively stained vesicle fluid or lesion material. Some pox viruses can be cultured on
the chorio-allantoic membrane of chick embryos, where they form pocks, and some can
be isolated by cell-culture.

HUMAN INFECTION

Vaccinia - a virus strain which has been used for immunisation against smallpox.
It's origins are not known but it seems to be a genetically distinct type of pox virus which
grows readily in a variety of hosts. In man it causes a localised pustule with scar
formation. In immunocompromised persons or eczematous persons it sometimes caused a
severe generalised vaccinia infection. Routine immunisation of all children in RSA
stopped in about mid 1980's. (Last case of smallpox in RSA was on Transvaal-Botswana
border in 1971.)

Cowpox - is acquired by humans usually by milking cows; it then manifests as

ulcerative lesions (sometimes called "milkers nodules") on the hands of dairy workers. It
was noted to protect against smallpox and was used by Jenner as a vaccine strain to
protect persons against smallpox. Despite its name, rodents are the main reservoir of
cowpox; it spreads secondarily to cows and domestic cats.

Molluscum contagiosum - is a minor infectious warty papule of the skin with a central
umbilication, transferred by direct contact, sometimes as a veneral disease.
Click here to see electron micrograph

Monkey pox - is a rare smallpox like disease of children in central Africa. It is acquired
from monkeys or wild squirrels, but does occasionally spread from man to man in
unvaccinated communities. Antigenically cross-reacts with other poxviruses. Sick
monkeys have not been identified, but apparently healthy animals have antibodies.

Pseudocowpox - occurs worldwide and is a disease primarily of cattle. In humans it

causes non-ulcerating "milker's nodes".
ORF - a worldwide occupational disease associated with handling sheep and goats
afflicted with "scabby mouth". In humans it manifests as a single painless, papulo-
vesicular lesion on the hand, forearm or face.

Historical Information regarding

SMALLPOX (the only disease that has been globally eradicated)

Source : only from close contact with infected persons. No animal reservoir or vector. No
latency in man.

Spread : by close contact, eg. within households, via droplet infection of pharyngeal
secretions (ulcers in pharynx).

Incubation : 10 - 12 days.

Major illness : Rather similar to severe chicken pox (varicella). Abrupt onset of fever
and prostration with a macular rash on the third day (head, limbs, hands and feet rather
than trunk, including palms and soles). Progressed to vesicles which become pustular,
ulcerated, scabbed, healed with scarring ("pock marked face"). Also inside mouth. 16 to
30% mortality. Milder subtypes were also described.

EPIDEMIOLOGY

As result of an unprecedented international WHO campaign of 'seek and contain' by

vaccination, the last natural case of smallpox was diagnosed in 1977. Despite intensive
searching, no other cases have been detected in 17 years. In 1994, smallpox virus remains
only in laboratory deepfreezers in Moscow and Atlanta, USA, awaiting a politico-
scientific decision for final extinction. It's genetic sequence information is stored in
computer memory banks.

POX VIRUSES AS VACCINE VECTORS

Certain poxvirus strains which cause localised lesions or abortive infections in mammals
(vaccinia) or birds (avipox) are currently being studied as possible recombinant vaccine
vectors for use in man and domestic animals.

Genes coding for immunising antigens of a variety of pathogens may be inserted into the
large poxvirus genome, and some undesirable genes of the pox virus may be excised. A
vaccinia-based rabies vaccine has been successfully used on wild foxes (via bait) in
Europe. Half-a-dozen others are in clinical trial.
 RABIES
INTRODUCTION
Rabies is an infectious disease of animals caused by a bullet-shaped, enveloped RNA
virus, 180 x 75 nm. Man is occasionally infected, and once infection is established in the
CNS, the outcome is almost invariably fatal.
HUMAN RABIES
Is acquired from virus in saliva entering a bite wound caused by an infected animal,
usually a rabid dog. The severity of the bite determines the risk of infection. The disease
does not usually spread from man to man.

Incubation
After inoculation, the virus enters small nerve endings at the site of the bite. The virus
slowly travels up the nerve to reach the CNS where it replicates and then travels down
nerves to the salivary glands where there is further replication. The time it takes to do this
depends upon the length of the nerve - a bite on the foot will have a very much longer
incubation period than a bite on the face. The incubation period may last from two
weeks to six months. Very often the primary wound is healed and forgotten by the time
of clinical presentation.

Clinical Presentation

A) Furious Rabies
When the virus reaches the CNS the patient presents with headache, fever, irritability,
restlessness and anxiety. This may progress to muscle pains, salivation and vomiting.
After a few days to a week the patient may experience a stage of excitement and be
wracked with painful muscle spasms, triggered sometimes by swallowing of saliva or
water. Hence they drool and learn to fear water (* Hydrophobia). The patients are also
excessively sensitive to air blown on the face. The stage of excitement lasts only a few
days before the patient lapses into coma and death.
Once clinical disease manifests, there is a rapid, relentless progression to invariable
death, despite all treatment.

B) Dumb Rabies
Starts in the same way, but instead of progressing into excitement, the subject retreats
steadily and quietly downhill, with some paralysis, to death. Rabies diagnosis may easily
be missed.

ANIMAL RABIES
Very similar picture to human rabies. In the stage of excitement the animal may bite
vigorously and viciously at anything: sticks, stones, grass, other animals and humans,
without provocation.

Wild animals may be abnormally tame or appear sick - beware of approaching or picking
up such an animal ("dumb rabies").
Epidemiology

1. The disease is endemic in wild animals in most parts of the world although some
countries (UK, Australia) are rabies free through vigorous control. The wild animal cycle
constitutes the natural reservoir.

2. Wild animals may bite and infect domestic animals (cattle, horses, pigs, dogs and
cats) which in turn may infect man. Occasionally wild animals may infect man directly.

3. In recent decades, a separate form of dog rabies (spread from dog to dog) has been
recognised as spreading from West Africa eastwards and southwards in Africa. Via
Mozambique, it reached Natal Kwazulu in the late 1970's and early 80's. Semi-wild dogs
in Natal have formed the highest endemic rabies reservoir and source for human cases in
the whole of the RSA.

ANIMAL RESERVOIR

• MONGOOSE (main reservoir in RSA in the wild)

• SURICATE
• JACKAL
• BAT (some evidence to suggest carrier status and droplet infection)
• FOX (in Europe)
• SKUNKS, RACCOONS (in USA)
• SEMI-WILD DOGS (in Natal)

Note: As in man, an infected animal becomes sick and dies. There is no substantial
evidence of a true carrier status in apparently well animals, except perhaps in bats.

DIAGNOSIS
By assessment of:
1.Bite
Geographical area, type of animal, severity and site of bite.

2. Animal

Live - observe in cage:

If survives > 8 days, then NOT rabies.
Dead - brain sent to Onderstepoort
- Negri bodies
- IFA
 - virus isolation

3. Man

Live - difficult diagnosis

- clinical picture, skin biopsy, corneal impression (antibodies only appear very
late)
Dead - brain sent to Onderstepoort
"Negri bodies" in cytoplasm of brain cells;
immunofluorescence virus isolation

TREATMENT

• 1. Wash wound (soap, detergent and water)

• 2. Anti-rabies serum (human). Passive immunisation.
• 3. Vaccine (intensive course). Active immunisation.

RABIES VACCINE
A good but expensive killed virus vaccine (Human Diploid Cell Vaccine, HDCV)
grown in human fibroblasts is available for safe use in man.
The unusually long incubation period of the virus permits the effective use of active
immunisation with vaccine post-exposure. When used, vaccine has dramatically cut the
rabies death rate. Supplied free by the State through district surgeons in South Africa.
(Older killed virus vaccines, made from infected neural tissues, were poorly
immunogenic and had allergic encephalitic side effects, but are still used in developing
countries.)

Prophylaxis
High-risk persons, eg. veterinarians, may be immunised before exposure, and then merely
require one or two booster doses if they should be exposed to rabies.

Animal Vaccines
A range of live and killed virus vaccines are available for domestic animals (farm
animals, cats and dogs).

Experimental vaccination of wildlife by using recombinant vaccinia vaccine (live) in

bait for foxes in Europe and North America has been quite promising.

CONTROL OF RABIES (Government Department of Veterinary Services)

1. Education
2. Vaccination of dogs, cats and farm animals.
3. Notification
- animals (district vet officer, police, magistrate) - human (district surgeon)
 HUMAN RETROVIRUSES
Retroviruses infect a wide range of animal species
and cause a variety of diseases including:
tumours, wasting and auto-immune diseases, immunodeficiency syndromes and aplastic
and haemolytic anaemias.

Human retroviruses
Four human reteroviruses have been identified. All infect CD4 bearing cells. These were
only identified in the 1980's when it became possible to culture T-cells in vitro.

• HTLV1 - T-cell leukaemias/lymphomas

Tropical spastic paraparesis
• HTLV2 - No known pathology
• HIV 1 & 2 - AIDS

They are enveloped viruses, with an RNA genome. The name is derived from the fact
that the virus particle contains an RNA-dependent DNA Polymerase (Reverse
transcriptase). This enzyme converts the RNA genome into DNA, which then integrates
into the host chromosomal DNA. The reverse transcriptase is highly error prone and rapid
genetic variation is a feature of this group.
The genus is divided into three sub families:
Oncovirinae - oncogenic - cause sarcomas and leukaemias in animals;
eg Rous Sarcoma Virus
Lentivirinae - slow progressive degenerative disorders
eg visnavirus of sheep, human immunodeficiency virus (HIV)
 Spumavirinae - no pathology known

Defective virus
Virus which has lost a gene that is essential for replication and can, therefore, only
undergo productive infection if the cell that is harbouring the virus is superinfected with a
helper virus that can supply the function of the lost gene.
Many oncogenic retroviruses are defective.

Endogenous reteroviral sequences

Defective retroviruses which integrate into the host genome and are passed down from
generation to generation. Two percent of the human genome is made up of endogenous
retroviral sequences.

Genome organization The virus has a diploid genome (2 copies of RNA genome per
virus particle). The genome codes for at least three genes: gag, pol and env.

• gag - codes for the core proteins, structural virion components

• pol - reverse transcriptase
• env - envelope glycoprotein

LTR -Long terminal repeat - is a regulatory sequence at each end of the genome.

LTR - gag - pol - env - (onc) - LTR

Their presence allows integration into the host chromosome and controls gene
expression

onc - (see oncogenes)

Life cycle of lentiviruses

 Oncogenes

Viruses of the Oncovirinae subfamily contain an extra gene termed, 'onc'. These genes
are called oncogenes because their expression in the virus infected cell is associated with
tumour production. These genes were originally acquired from the host cell itself. The
cellular gene was picked up during integration of the virus genome into the host DNA,
way back in evolution. Most oncogenes code for proteins with growth promoting
properties and their expression can lead to uncontrolled proliferation of the infected cell
and tumour development.

HTLV 1 & 2
Human T-Cell Lymphotrophic Viruses, belong to the sub-family Oncovirinae.

HTLV1

Clinical features
1. T-cell leukaemia/lymphoma. Aggressive tumour of CD4 cells which infiltrates skin
and brain. Tumours are only produced after a prolonged latent period. The virus does not
contain an oncogene and it is believed that the malignant change is the result of
interruption and disregulation of host DNA, by viral genome insertion. Less than 1% of
HTLV 1 infected individuals develop this malignancy.

2. Tropical spastic paraparesis. Aggressive non-demyelinating spastic paraparesis.

Epidemiology
Seroprevalence is high in South West Japan, the Caribbean and parts of West Africa. In
high incidence areas up to 10% of adults may be infected. The seroprevalence increases
with age and family clustering of infection is common. Spread occurs through blood
transfusion and sexual intercourse. Mother to child transmission through breast feeding
has also been shown.
Laboratory Diagnosis
HTLV 1 specific antibody, ELISA.

HTLV 2

The virus shares extensive nucleic acid homology with HTLV 1. It was first isolated from
a patient with hairy cell leukaemia, but no specific pathology has been attributed to it.

HIV 1 and 2
Human Immunodeficiency Viruses

Background:-
In the spring of 1981, previously healthy homosexual men in New York and Los Angeles
were found to be suffering from severe immunodeficiency states associated with severe
opportunistic infections and rare malignancies.

At this time an infectious aetiology was suspected especially when it was perceived the
disease could be transmitted by blood transfusions and blood products. (Haemophiliacs)

In 1983, a new retrovirus termed LAV (now called HIV 1) was isolated from the T-cells
of a patient with persistent generalised lymphadenopathy.

In 1986, a second closely related virus, termed HIV 2 was isolated from a patient from
West Africa with acquired immunodeficiency syndrome, AIDS.

Currently, about 14 million people are believed to be infected, world-wide; nine million
of these are in Africa (see figure 2). HIV1 is the major cause of the AIDS pandemic;
HIV2 is of lower virulence and infection has largely remained confined to West Africa.
Origin
AIDS appears to be a new disease. The origin of the virus is obscure but it is probable
that infection arose in Africa. Retrospective screening of stored sera has shown that the
earliest known infection occurred in Kinshasa, Zaire, in 1959.
HIV is believed to have evolved from related viruses that infect African monkeys,
namely the simian immunodeficiency viruses (SIV) (which cause a similar disease to
AIDS in monkeys). HIV 2 is more closely related to SIV than HIV 1. It is not known how
the original transfer of virus from monkey to man took place. These viruses mutate
readily which explains the rapid subsequent divergence.(figure 4)

Spread(figure 3)

Epidemiology - South Africa

HIV was introduced into South Africa from two sources:
a) Into the homo-sexual community - from North America and Europe,and
b) into the hetero-sexual community - from north of our borders by truck drivers,
returning exiles, migrant labourers.

A survey, conducted in October/November 1993, of women attending ante natal clinics in

different parts of the country has shown that the prevalence of HIV infection varies from
1.33% in the Cape to 9.62% in KwaZulu/Natal.

Transmission
Infection is transmitted in a manner identical to that of hepatitis B.
1.) Blood products

• Blood transfusions
• Intra-venous drug abusers - sharing of needles
• Health care workers:
needlestick injuries - risk approximately 0.36% (depends on extent of the injury)
muco-cutaneous exposure - no sero-conversion incidents have been reported

2.) Organ transplants

3.) Sexual intercourse
Both homosexual and hetero-sexual exposure
Increased risk of transmission if partners have other sexually transmitted diseases

4.) Vertical Transmission

10-40% of babies born of HIV-infected mothers will be infected.

Infection may occur

in utero
during birth
post-natally, through breast feeding
NB! THERE IS NO EVIDENCE TO SUGGEST THAT HIV CAN BE
TRANSMITTED BY:
 Insects
Casual contact saliva, kissing
sharing of eating and drinking utensils

Clinical features

Primary infection
About 90% of patients develop a flu-like illness which co-incides with seroconversion,
between 2 and 4 weeks post exposure. Symptoms include, fever, night sweats, sore
throat, lymphadenopathy, diarrhoea. The illness is self limiting.

Asymptomatic phase
Of variable duration, from 2 to 10 years. Patients are clinically well, but infectious.

Prodromal phase
This period is heralded by the insidious onset of a variety of prodromal disorders,
including: weight loss, fever, persistant lymphadenopathy, oral candidiasis and diarrhoea.
These symptoms precede the progression to AIDS.

Acquired Immunodeficiency Syndrome (AIDS)

Syndrome with the following features:

1) Constitutional disease: fever, diarrhoea, weight loss, skin rashes

2) Neurological disease: dementia, myelopathy, peripheral neuropathy

3) Immunodeficiency: Increased susceptibility to opportunistic infections: (see
table 1)
4) Rare malignancies: Kaposi sarcoma, oral hairy leukoplakia, lymphomas.

Kaposi sarcoma - is a tumour of endothelial cells. Prior to the AIDS epidemic, this
tumuor was rare and only found in middle aged African and Mediterranean Jewish men,
in whom it was an indolent condition. AIDS patients develop a disseminated highly
aggressive form of the disease.

Paediatric Infection
Following infection in the perinatal period, babies may develop a progressive illness in
the first few months of life (No latent period). Clinical features include: Failure to thrive,
diarrhoea, lymphadenopathy, susceptibility to opportunistic infections hepato-
splenomegaly, lymphoid interstitial pneumonia and parotitis.

Pathogenesis
 HIV infects CD4 cells

Disseminated infection

Specific immune Response

Antibody
Cell mediated immunity

Clearance of most virus

Some persistence
a) Gradual loss of CD4 cells
b) Destruction of microenvironment of lymphoid tissue

IMMUNO-DEFICIENCY

Cell tropism: CD4+ T cells, Macrophages

Destruction of CD4 cells occurs through the following mechanisms:

direct lysis of infected cells
syncytium formation

Infection of precursor cells and destruction of microenvironment

Impairment of CD4 cell function

LABORATORY DIAGNOSIS

Serology
IgG develops 4-6 weeks post exposure and remains detectable for life. Its presence in
serum therefore indicates infection.
Exception: Uninfected infants of HIV positive mothers

Direct detection of virus

p24 antigen ELISA
culture from PBMC's
PCR

VACCINE PROSPECTS
There is no effective vaccine available for HIV. Attempts have been made to develop a
vaccine, using:

• purified viral envelope glycoproteins, gp120 or 160

• whole inactivated virus
• live attenuated HIV strains (lacking certain genes)
• live recombinant virus vectors, expressing HIV proteins.

A major difficulty is the fact that neutralizing antibody in the serum does not protect the
host from infection with HIV: Possible reasons for this include:

• Antibody enhancement of infection

• Rapid virus mutation may result in variation of envelope antigens (escape
mutants)
• HIV can infect cells in sites that are sequestered from antibody
• Host may be infected by whole virus-infected cells

ANTI-VIRAL THERAPY

ANTI-VIRAL CHEMOTHERAPY

Antiviral Agents

Antiviral drugs are available to treat only a few viral diseases. The reason for this is the
fact that viral replication is so intimately associated with the host cell that any drug that
interferes significantly with viral replication, is likely to be toxic to the host.

Two useful antivirals are:

the nucleoside analogues and
the interferons
but there are other targets in the different stages of intracellular viral growth which show
potential for antiviral chemotherapy.

Stages in virus replication which are possible targets for chemotherapeutic

agents:

• Attachment to host cell

• Uncoating - (Amantadine)
• Synthesis of viral mRNA - (Interferon)
• Translation of mRNA - (Interferon)
 • Replication of viral RNA or DNA - (Nucleoside anologues)
• Maturation of new virus proteins (Protease inhibitors)
• Budding, release

Diseases for which effective therapy is available:

Herpes Simplex virus (Acyclovir)

Varicella-Zoster virus (Acyclovir)
Cytomegalovirus (Gancyclovir, Foscarnet)
AIDS (Zidovudine, Lamivudine[3TC], Protease inhibitors; in combination)
Respiratory Syncitial virus (Ribavirin)
Influenza (Amantadine)

Nucleotide analogues:

These are synthetic compounds which resemble nucleosides, but have an incomplete or
abnormal deoxy-ribose /or ribose group.
These compounds are phosphorylated to the tri-phosphate form within the infected cell.
In this form, the drug competes with normal nucleotides for incorporation into viral
DNA or RNA. Incorporation into the growing nucleic acid chain results in irreversible
association with the viral polymerase and chain termination.
 Interferons:

There are three classes: alpha- beta- and gamma-

The alpha and beta Interferons

are cytokines which are secreted by virus infected cells.
They bind to specific receptors on adjacent cells and protect them from infection by
viruses.
They form part of the immediate protective host response to invasion by viruses.
In addition to these direct antiviral effects, alpha and beta interferon also enhance the
expression of class I and class II MHC molecules on the surface of infected cells, in
this way, enhancing the presentation of viral antigens to specific immune cells. Their
presence can be demonstrated in body fluids during the acute phase of virus infection.

Recombinant alpha and beta interferons are now available and have been used for the
treatment of
Chronic hepatitis B and C virus infections.

However, side effects such as fever, malaise and weight loss have limited the use.

gamma Interferon (immune interferon)

is a cytokine secreted by TH1 CD4 cells.
Its function is to enhance specific T cell mediated immune responses.

Mechanism of action of the interferons :

1. Enhancement of the specific immune response.

By increasing the expression of MHC class I molecules on the surface of infected
cells, the interferons increase the opportunity for specific cytotoxic T cells to
recognise and kill infected cells.
2. Direct antiviral effect
a) degradation of viral mRNA
b) inhibition of protein synthesis
Prevents the infection of new cells

Immunoglobulin Therapy

Passive Immunisation

Passive immunisation is the transfer of immunity to a host by means of

immunoglobulins (preformed antibodies). These immunoglobulins are typically prepared
by cold ethanol fractionation as a 16% solution of gammaglobulin from large pools of
serum obtained from the blood donations of at least 1000 donors.

Immunoglobulin from immune individuals can be used as prophylaxis to prevent viral

infections in exposed, but non immune individuals. It works by binding to extra-cellular
virions and preventing them from attaching to and entering susceptible cells. The
protective effect is short lived (up to three months) because the antibodies are
metabolised by the host.

"Normal" Immune globulin

This is a pooled product, prepared from the serum of normal blood donors. It contains
low titres of antibody to a wide range of human viruses. It is mainly used as prophylaxis
against:

• hepatitis A virus infection,

• parvovirus infection, and
• enterovirus infections (in neonates).
• It may also be given as passive protection to HIV-infected babies.

Hyper-immune globulin
Immunoglobulin may be prepared from the serum of selected individuals who have high
titres of antibody to particular viruses.
Examples include:

• Zoster immune globulin

Prevention of Varicella in immunocompromised children and neonates.
• Human Rabies immunoglobulin
Post-exposure prophylaxis in an individual who has been bitten by a rabid animal.
• Hepatitis B Immune globulin
Non-immune individual who has been exposed to HBV.
• RSV Immune globulin
Treatment of respiratory syncitial virus infections in the very young

VACCINES
The introduction of vaccination has been one of the most
decisive advances leading to the dramatic downward trend in
the incidence of many viral diseases.

The principle of vaccination is to induce a "primed" state in the vaccinated subject so

that, following exposure to a pathogen, a rapid secondary immune response is generated
leading to the accelerated elimination of the organism and protection from clinical
disease. Success depends on the generation of memory T and B cells and the presence in
the serum of neutralizing antibody.
Attributes of a good vaccine
1.Ability to elicit the appropriate immune response for the particular pathogen:
Tuberculosis - cell mediated response
most bacterial and viral infections - antibody
2. Long term protection
ideally life-long
3. Safety
vaccine itself should not cause disease
4. Stable
retain immunogenicity, despite adverse storage conditions prior to
administration
5. Inexpensive

Types of Vaccine

Vaccines in general use include: LIVE vaccines; and KILLED vaccines

Vaccines are available for:

• Hepatitis B virus
• Hepatitis A virus
• Influenza
• Measles
• Mumps
• Polio
• Rubella
• Rabies
• Yellow Fever
• Varicella Zoster
 A. Live Vaccines

1. Live attenuated organisms

Organisms whose virulence has been artificially reduced by in vitro culture under
adverse conditions, such as reduced temperature. This results in the selection of mutants
which replicate poorly in the human host and are therefore of reduced
virulence. Replication of the vaccine strain in the host reproduces many of the features of
wild type infection, without causing clinical disease. Most successful viral vaccines
belong to this group.

The immune response is usually good - when the virus replicates in the host cells, both
antibody as well as cell mediated immune responses are generated and immunity is
generally long lived. Often, only a single dose is needed to induce long term immunity.

Potential drawbacks to these vaccines include:

the danger of reversion to virulence and
the possibility of causing extensive disease in immunocompromised individuals.

2. Heterologous vaccines

Closely related organism of lesser virulence, which shares many antigens with the
virulent organism. The vaccine strain replicates in the host and induces an immune
response that cross reacts with antigens of the virulent organism. The most famous
example of this type of vaccine is vaccinia virus: Both cowpox virus and vaccinia virus
are closely related to variola virus, the causitive agent of smallpox. The eighteenth
centuary physician, Edward Jenner observed that milkmaids who had been infected with
cowpox virus were immune to smallpox. Widespread use of vaccinia virus as a vaccine
has lead to the world-wide eradication of smallpox.

3. Live recombinant vaccines

It is possible, using genetic engineering, to introduce a gene coding for an immunogenic

protein from one organism into the genome of another (such as vaccinia virus). The
organism expressing a foreign gene is called a recombinant. Following injection into the
subject, the recombinant organism will replicate and express sufficient amounts of the
foreign protein to induce a specific immune response to the protein.

Attributes - live vaccines

Good immune response

• Both Cell Mediated Immunity and antibody responses.

• Immunity is long lived
• Single dose
Safety

• Danger of reversion to virulence, or

• Severe disease in immunocomprised

Stability

• Organisms in the vaccine must remain viable in order to infect and replicate in the
host
• Vaccine preparations are therefore very sensitive to adverse storage conditions
• Maintenance of the cold chain is very important.

Expense

• Cheap to prepare

Killed (inactivated) vaccines

When safe live vaccines are not available, either because attenuated strains have not been
developed or else because reversion to wild type occurs too readily, it may be possible to
use an inactivated preparation of the virulent organism to immunize the host.

The organism is propagated in bulk, in vitro, and inactivated with either beta-
propiolactone or formaldehyde. These vaccines are not infectious and are therefore
relatively safe. However, they are usually of lower immunogenicity and multiple doses
may be needed to induce immunity. In addition, they are usually expensive to prepare.

Subcellular fractions

When protective immunity is known to be directed against only one or two proteins of an
organism, it may be possible to use a purified preparation of these proteins as a vaccine.
The organism is grown in bulk and inactivated, and then the protein of interest is purified
and concentrated from the culture suspension. These vaccines are safe and fewer local
reactions occur at the injection site. However, the same disadvantages of poor
immunogenicity and the need for multiple boosters applies.

Recombinant proteins

Immunogenic proteins of virulent organisms may be synthesized artificially by

introducing the gene coding for the protein into an expression vector, such as E-coli or
yeasts. The protein of interest can be extracted from lysates of the expression vector, then
concentrated and purified for use as a vaccine. The only example of such a vaccine, in
current use, is the hepatitis B vaccine.
Attributes - Killed vaccines

Immune response

• poor; only antibody - no cell immediated immune response.

• response is short-lived and multiple doses are needed.
• may be enhanced by the incorporation of adjuvants into the vaccine preparation
(see below)

1. Safety

• Inactivated, therefore cannot replicate in the host and cause disease.

• Local reactions at the site of injection may occur.

2. Stability

• Efficacy of the vaccine does not rely on the viability of the organisms.
• These vaccines tend to be able to withstand more adverse storage conditions.

3. Expense

• Expensive to prepare.

Adjuvants

Certain substances, when administered simultaneously with a specific antigen, will

enhance the immune response to that antigen. Such compounds are routinely included in
inactivated or purified antigen vaccines.

Adjuvants in common use:

1. Aluminium salts

• First safe and effective compound to be used in human vaccines.

 • It promotes a good antibody response, but poor cell mediated immunity.

2. Liposomes and Immunostimulating complexes (ISCOMS)

3. Complete Freunds adjuvant is an emulsion of Mycobacteria, oil and water

• Too toxic for man

• Induces a good cell mediated immune response.

4. Incomplete Freund's adjuvant as above, but without Mycobacteria.

5. Muramyl di-peptide

• Derived from Mycobacterial cell wall.

6. Cytokines

• IL-2, IL-12 and Interferon-gamma.

Possible modes of action:

• By trapping antigen in the tissues, thus allowing maximal exposure to dendritic

cells and specific T and B lymphocytes.
• By activating antigen-presenting cells to secrete cytokines that enhance the
recruitment of antigen-specific T and B cells to the site of inoculation.

DNA Vaccines
DNA vaccines are at present experimental, but hold promise for future therapy since they
will evoke both humoral and cell-mediated immunity, without the dangers associated
with live virus vaccines.

The gene for an antigenic determinant of a pathogenic organism is inserted into a

plasmid. This genetically engineered plasmid comprises the DNA vaccine which is then
injected into the host. Within the host cells, the foreign gene can be expressed
(transcribed and translated) from the plasmid DNA, and if sufficient amounts of the
foreign protein are produced, they will elicit an immune response.
 Vaccines in general use

Measles

Live attenuated virus grown in chick embryo fibroblasts, first introduced in the 1960's. Its
extensive use has led to the virtual eradication of measles in the first world. In developed
countries, the vaccine is administered to all children in the second year of life (at about
15 months). However, in developing countries, where measles is still widespread,
children tend to become infected early (in the first year), which frequently results in
severe disease. It is therefore important to administer the vaccine as early as possible
(between six months and a year). If the vaccine is administered too early, however, there
is a poor take rate due to the interference by maternal antibody. For this reason, when
vaccine is administered before the age of one year, a booster dose is recommended at 15
months.

Mumps

Live attenuated virus developed in the 1960's. In first world countries it is administered
together with measles and rubella at 15 months in the MMR vaccine. This is not a legal
requirement in South Africa.

Rubella

Live attenuated virus. Rubella causes a mild febrile illness in children, but if infection
occurs during pregnancy, the foetus may develop severe congenital abnormalities. Two
vaccination policies have been adopted in the first world. In the USA, the vaccine is
administered to all children in their second year of life (in an attempt to eradicate
infection), while in Britain, until recently, only post pubertal girls were vaccinated. It
was feared that if the prevalence of rubella in the community fell, then infection in the
unimmunized might occur later - thus increasing the likelihood of infection occurring in
the child-bearing years. This programme has since been abandoned in Britain and
immunization of all children is the current practice.

Polio

Two highly effective vaccines containing all 3 strains of poliovirus are in general use:

• The killed virus vaccine (Salk, 1954) is used mainly in Sweden, Finland,
Holland and Iceland.
• The live attenuated oral polio vaccine (Sabin, 1957) has been adopted in most
parts of the world; its chief advantages being: low cost, the fact that it induces
 mucosal immunity and the possibility that, in poorly immunized communities,
vaccine strains might replace circulating wild strains and improve herd
immunity. Against this is the risk of reversion to virulence (especially of types 2
and 3) and the fact that the vaccine is sensitive to storage under adverse
conditions.
• The inactivated Salk vaccine is recommended for children who are
immunosuppressed.

Hepatitis B

Two vaccines are in current use: a serum derived vaccine and a recombinant vaccine.
Both contain purified preparations of the hepatitis B surface protein.

The serum derived vaccine is prepared from hepatitis B surface protein, purified from the
serum of hepatitis B carriers. This protein is synthesised in vast excess by infected
hepatocytes and secreted into the blood of infected individuals. A vaccine trial performed
on homosexual men in the USA has shown that, following three intra-muscular doses at
0, 1 and 6 months, the vaccine is at least 95% protective.

A second vaccine, produced by recombinant DNA technology, has since become

available. Previously, vaccine administration was restricted to individuals who were at
high risk of exposure to hepatitis B, namely: infants of hepatitis B carrier mothers, health
care workers, homosexual men and intravenous drug abusers. However, hepatitis B has
been targetted for eradication , and since 1995 the vaccine has been included in the
universal childhood immunization schedule. Three doses are given; at 6, 10, and 14
weeks of age. As with any killed viral vaccines, a booster will be required at some
interval (not yet determined, but about 5 years) to provide protection in later life from
hepatitis B infection as a venereal disease.

Hepatitis A

A vaccine for hepatitis A has been developed from formalin-inactivated , cell culture-
derived virus.
Two doses, administered one month apart, appear to induce high levels of neutralising
antibodies. The vaccine is recommended for travellers to third world countries, and
indeed all adults who are not immune to hepatitis A.

Yellow Fever

The 17D strain is a live attenuated vaccine developed in 1937. It is a highly effective
vaccine which is administered to residents in the tropics and travellers to endemic areas.
A single dose induces protective immunity to travellers and booster doses, every 10
years, are recommended for residents in endemic areas.

Rabies
No safe attenuated strain of rabies virus has yet been developed for humans. Vaccines in
current use include:

• The neurotissue vaccine - here the virus is grown in the spinal cords of rabbits,
and then inactivated with beta-propiolactone. There is a high incidence of
neurological complications following administration of this vaccine due to a
hypersensitivity reaction to the myelin in the preparation and largely it has been
replaced by
• A human diploid cell culture-derived vaccine (also inactivated) which is much
safer.

There are two situations where vaccine is given:

a) Post-exposure prophylaxis, following the bite of a rabid animal:
A course of 5-6 intramuscular injections, starting on the day of exposure. Hyperimmune
rabies globulin may also administered on the day of exposure.

b) Pre-exposure prophylaxis is used for protection of those whose occupation puts them
at risk of infection with rabies; for example, vets, abbatoir and laboratory workers.
This schedule is 2 doses one month apart ,and a booster dose one year later. (Further
boosters every 2-3 years should be given if risk of exposure continues).

Influenza

Repeated infections with influenza virus are common due to rapid antigenic variation of
the viral envelope glycoproteins. Antibodies to the viral neuraminidase and
haemagglutinin proteins protect the host from infection. However, because of the
rapid antigenic variation, new vaccines, containing antigens derived from influenza
strains currently circulating in the community, are produced every year.
Surveillance of influenza strains now allows the inclusion of appropriate antigens for
each season.The vaccines consist of partially purified envelope proteins of inactivated
current influenza A and B strains.

Individuals who are at risk of developing severe, life threatening disease if infected with
influenza should receive vaccine. People at risk include the elderly,
immunocompromised individuals, and patients with cardiac disease. In these patients,
protection from disease is only partial, but the severity of infection is reduced.

Varicella-Zoster virus

A live attenuated strain of varicella zoster virus has been developed. It is not licensed in
South Africa for general use, but is used in some oncology units to protect immuno-
compromised children who have not been exposed to wild-type varicella zoster virus.
Such patients may develop severe, life threatening infections if infected with the wild
type virus.
 An illustrated tutorial.

Laboratory diagnosis of viral infections frequently requires the isolation of the virus in
cell cultures. Cell monolayers are inoculated with a suitable clinical specimen, and then
observed for cytological changes that indicate virus growth.

Cytopathic changes

The term "cytopathic effect" (CPE) is frequently applied to virus-induced cellular changes
that are visible by light microscopy. These changes include swelling or shrinkage of cells,
the formation of multinucleated giant cells (syncytia), and the production of "inclusions"
(made visible by staining) in the nucleus or cytoplasm of the infected cell.

The most efficient way to demonstrate cellular changes is by staining with chromatic
dyes.
Cell monolayers are fixed and then exposed to basic and acidic dyes that accentuate the
nature and location of the changes.
 The use of haemotoxylin (basic dye) and eosin (acidic dye) is often referred to as
H&E staining.

The gross appearance of the cellular changes, and the location and nature of the
"inclusions" - i.e. basophilic or eosinophilic - can in many instances be used as a
diagnostic criterion to identify the causative virus. These will be illustrated for some of
the viruses commonly isolated in cell culture:-

• Adenovirus - example of basophilic intranuclear inclusions

•
•
• Reovirus - example of eosinophilic intra-cytoplasmic inclusions

See also
Syncytia: The herpes group of viruses:
Measles virus herpes simplex virus
Respiratory syncytial virus human cytomegalovirus
Mumps virus varicella zoster virus

Adenovirus

Cells infected with adenovirus have an

affinity for haematoxylin (a purplish-blue
dye).
Infected cells become rounded and the cell
sheet disintegrates. Dark basophilic
inclusions within the nuclei represent
accumulated viral proteins at the site of virus
assembly.
Reovirus
Replication of reovirus particles occurs in the cytoplasm of the cell, and in the final
stages of assembly the virus particles bud through the endoplasmic reticulum membrane.
Cytoplasmic sites of accumulated viral protein are stained with eosin (deep pink).