Avian Influenza(Bird Flu)

Md. Shaifur Rahman

Biotechnology and Genetic Engineering Discipline
Khulna University
Khulna-9208
Bangladesh
Email: mdshaifur@gmail.com
 Introduction
Avian influenza is an infection caused by avian (bird) influenza (flu) viruses.
These influenza viruses occur naturally among birds. Wild birds carry the
viruses in their intestines, but usually do not get sick from them. However, avian
influenza is very contagious among birds and can make some domesticated
birds, including chickens, ducks and turkeys, very sick and kill them. Infected
birds shed influenza virus in their saliva, nasal secretions, and feces.
Domesticated birds may become infected with avian influenza virus through
direct contact with infected waterfowl or other infected poultry, or through
contact with surfaces (such as dirt or cages) or materials (such as water or
feed) that have been contaminated with the virus.

Infection with AIV in domestic poultry causes two main forms of disease that are
distinguished by low and high extremes of virulence. The “low pathogenic”
usually causes only mild symptoms (such as ruffled feathers and a drop in egg
production). However, the highly pathogenic form spreads more rapidly through
flocks of poultry which cause disease that affects multiple internal organs and
has a mortality rate that can reach 90-100% often within 48 hours. The risk from
avian influenza is generally low to most people, because the viruses do not
usually infect humans
 Historical Background
‰ H5N1, Hong Kong, Special Administrative Region, 1997: Highly pathogenic
avian influenza A (H5N1) infections occurred in both poultry and humans. This was
the first time an avian influenza A virus directly transmitted from birds to humans.
‰ H9N2, China and Hong Kong, 1999: Low pathogenic avian influenza A (H9N2)
virus infection was confirmed.
‰ H7N2, Virginia, 2002: one person was found to have serologic evidence of
infection with H7N2.
‰ H7N7, Netherlands, 2003: 89 people were confirmed to have H7N7 influenza
virus infection associated with this poultry outbreak. These cases occurred mostly
among poultry workers.
‰ H7N3, Canada, 2004: human infections of highly pathogenic avian influenza A
(H7N3) among poultry workers were found.

‰H5N1, Cambodia, China, Indonesia, Thailand and Vietnam, 2005:

‰H5N1, Azerbaijan, Cambodia, China, Djibouti, Egypt, Indonesia, Iraq,

Thailand, Turkey, 2006:

‰H5N1, India, Bangladesh,2007

 Objectives

The recent outbreaks of the Highly Pathogenic Avian Influenza viruses

among poultry in many countries, with human infection and deaths,
have prompted the World Health Organization to strongly recommend
its member states to rapidly prepare themselves in dealing with
influenza pandemic. Therefore, it is necessary for Bangladesh to take
the necessary steps against Influenza pandemic.

The major objectives of this study are as follows:

1. To know the physical and genetical characteristics of
Avian Influenza Virus (AIV)
2. To prevent the outbreak of an influenza pandemic.
3. To reduce the morbidity and mortality from influenza.
4. Make a national strategic plan to face Influenza
pandemic.
 Types, Subtypes, and Strains

I. Influenza Type A and Its Subtypes

™ Influenza A (H5N1) virus – also called “H5N1 virus” – is

an influenza A virus subtype that occurs mainly in birds, is
highly contagious among birds, pigs, horses, and other
animals and can be deadly to them.

™ Influenza type A viruses are divided into subtypes and

named on the basis of two proteins on the surface of the
virus: hem agglutinin (HA) and neuraminidase(NA).

™ For example, an “H7N2 virus” designates influenza a

subtype that has an HA 7 protein and an NA 2 protein.
Similarly an “H5N1” virus has an HA 5 protein and an NA
1 protein.
™There are 16 known HA subtypes and 9 known NA
subtypes.
™Many different combinations of HA and NA proteins are
possible. Only some influenza A subtypes (i.e., H1N1, H1N2,
and H3N2) are currently in general circulation among people.
™Other subtypes are found most commonly in other animal
species. For example, H7N7, which has unusual zoonotic
potential and killed one person. H3N8 viruses cause illness in
horses, and H3N8 also has recently been shown to cause
illness in dogs. H5N1 virus does not usually infect people, but
infections with these viruses have occurred in humans. Most of
these cases have resulted from people having direct or close
contact with H5N1-infected poultry or H5N1-contaminated
surfaces.
Three prominent subtypes of the avian influenza A virus that are known to infect
both birds and people are:

i. Influenza A H5
™Nine potential subtypes of H5 are known and can be highly pathogenic or low
pathogenic.

™H5 infections, such as HPAI H5N1 viruses have been documented among
humans and sometimes cause severe illness or death.

ii. Influenza A H7
™Nine potential subtypes of H7 are known. H7 infection in humans is rare but can
occur among persons who have direct contact with infected birds.

™7 viruses have been associated with both LPAI (e.g., H7N2, H7N7) and HPAI
(e.g., H7N3, H7N7), and have caused mild to severe and fatal illness in humans.
iii. Influenza A H9
™Nine potential subtypes of H9 are known; influenza A H9 has
rarely been reported to infect humans (At least three H9 infections in
humans have been confirmed).

™However, this subtype has been documented only in a low

pathogenic form.

II. Influenza Type B

™Influenza B viruses are usually found only in humans.
™ These viruses are not classified according to subtype.
™ Influenza B viruses can cause morbidity and mortality among
humans, but in general are associated with less severe than
influenza A viruses.

III. Influenza Type C

™Influenza type C viruses cause mild illness in humans.
™These viruses are not classified according to subtype.
IV. Strains

™Influenza B viruses and subtypes of influenza A virus are further

characterized into strains.

™There are many different strains of influenza B viruses and of

influenza A subtypes.

™New strains of influenza viruses appear and replace older strains.

™This process occurs through antigenic drift. When a new strain of

human influenza virus emerges, antibody protection that may have
developed after infection or vaccination with an older strain may not
provide protection against the new strain.

™ Therefore, the influenza vaccine is updated on a yearly basis to

keep up with the changes in influenza viruses.
How Influenza Viruses Change: Drift and Shift

™Influenza viruses are dynamic and are continuously

evolving.

™Influenza viruses can change in two different ways:

antigenic drift and antigenic shift. Influenza viruses are
changing by antigenic drift all the time, but antigenic shift
happens only occasionally.

™Influenza type A viruses undergo both kinds of changes;

influenza type B viruses change only by the more gradual
process of antigenic drift.
 ANTIGENIC DRIFT

™Antigenic drift refers to small, gradual changes that occur through point
mutations in the two genes that contain the genetic material to produce the main
surface proteins, hemagglutinin, and neuraminidase.

™These point mutations occur unpredictably and result in minor changes to these
surface proteins.

™ Antigenic drift produces new virus strains that may not be recognized by
antibodies to earlier influenza strains.

™This process works as follows: a person infected with a particular influenza

virus strain develops antibody against that strain. As newer virus strains appear,
the antibodies against the older strains might not recognize the "newer" virus, and
infection with a new strain can occur. This is one of the main reasons why people
can become infected with influenza viruses more than one time and why global
surveillance is critical in order to monitor the evolution of human influenza virus
stains for selection of which strains should be included in the annual production
of influenza vaccine. In most years, one or two of the three virus strains in the
influenza vaccine are updated to keep up with the changes in the circulating
influenza viruses. For this reason, people who want to be immunized against
influenza need to be vaccinated every year.
 ANTIGENIC SHIFTS

™Antigenic shift refers to an abrupt, major change to produce

a novel influenza A virus subtype in humans that was not
currently circulating among people.

™Antigenic shift can occur either through direct animal

(poultry)-to-human transmission or through mixing of human
influenza A and animal influenza A virus genes to create a
new human influenza A subtype virus through a process
called genetic reassortment. Antigenic shift results in a new
human influenza A subtype.
Genetics and diversity of the viral strains
Genetic structure and related subtypes

Fig. The H in H5N1 stands Fig. The N in H5N1

for "Hemagglutinin", as stands for
depicted in this molecular "Neuraminidase", as
model. depicted in this ribbon
diagram.
™ H5N1 is a subtype of the species Influenza A virus of the Influenzavirus A genus
of the Orthomyxoviridae family.
™ The H5N1 subtype is an RNA virus.
™ It has a segmented genome of eight abbreviated as PB2, PB1, PA, HA, NP, NA,
M and NS.
™ HA codes for hemagglutinin, an antigenic glycoprotein found on the surface of the
influenza viruses and are responsible for binding the virus to the cell that is being
infected.
™ NA codes for neuraminidase, an antigenic glycosylated enzyme found on the surface
of the influenza viruses. It facilitates the release of progeny viruses from infected
cells (Couch, R.1996).
™ HA and NA are also used as the basis for the naming of the different subtypes of
influenza A viruses. This is where the H and N come from in H5N1.
™ The hemagglutinin (HA) and neuraminidase (NA) RNA strands specify the structure
of proteins.
 PROPERTIES of H5N1

Infectivity

™H5N1 is easily transmissible between birds facilitating a potential

global spread of H5N1. While H5N1 undergoes specific mutations
and reassorting creating variations which can infect species not
previously known to carry the virus, not all of these variant forms can
infect humans.

™H5N1 as an avian virus preferentially binds to a type of galactose

receptors that populate the avian respiratory tract from the nose to the
lungs and are virtually absent in humans, occurring only in and
around the alveoli, structures deep in the lungs where oxygen is
passed to the blood. Therefore, the virus is not easily expelled by
coughing and sneezing, the usual route of transmission (Shinya K.et.
al., 2006 and Van Riel D. et. al., 2006).
Virulence

™H5N1 has mutated into a variety of strains with differing pathogenic profiles,
some pathogenic to one species but not others, some pathogenic to multiple
species. Each specific known genetic variation is traceable to a virus isolate of a
specific case of infection.

™Through antigenic drift, H5N1 has mutated into dozens of highly pathogenic
varieties divided into genetic clades which are known from specific isolates, but
all currently belonging to genotype Z of avian influenza virus H5N1, now the
dominant genotype (Kou Z.et. al., 2005). H5N1 isolates found in Hong Kong in
1997 and 2001 were not consistently transmitted efficiently among birds and did
not cause significant disease in these animals.

™ In 2002 new isolates of H5N1 were appearing within the bird population of
Hong Kong. These new isolates caused acute disease, including severe
neurological dysfunction and death in ducks. This was the first reported case of
lethal influenza virus infection in wild aquatic birds since 1961(Sturm-Ramirez
K.M.et. al.,2004). Genotype Z emerged in 2002 through reassortment from
earlier highly pathogenic genotypes of H5N1 that first infected birds in China in
1996, and first infected humans in Hong Kong in 1997.
 Transmission and host range

™Infected birds transmit H5N1 through their saliva, nasal secretions, feces and blood.
Other animals may become infected with the virus through direct contact with these bodily
fluids or through contact with surfaces contaminated with them. H5N1 remains infectious
after over 30 days at 0 °C (32.0 °F) over one month at freezing temperature or 6 days at 37
°C (98.6 °F) and one week at human body temperature. So at ordinary temperatures it
lasts in the environment for weeks. In arctic temperatures, it doesn't degrade at all.

Fig.1.1 Influenza A virus, the virus that causes Avian flu. Transmission electron micrograph of negatively
stained virus particles in late passage.
(Source: Dr. Erskine Palmer, Centers for Disease Control and Prevention Public Health Image Library)
High mutation rate

™Influenza viruses have a relatively high mutation rate that is characteristic of

RNA viruses. The segmentation of the influenza genome facilitates genetic
recombination by segment reassortment in hosts who are infected with two
different influenza viruses at the same time (Kou Z. et. al., 2005). H5N1 viruses
can reassort genes with other strains that co-infect a host organism, such as a
pig, bird, or human, and mutate into a form that can pass easily among humans.

™The ability of various influenza strains to show species-selectivity is largely

due to variation in the hemagglutinin genes. Genetic mutations in the
hemagglutinin gene that cause single amino acid substitutions can significantly
alter the ability of viral hemagglutinin proteins to bind to receptors on the
surface of host cells. Such mutations in avian H5N1 viruses can change virus
strains from being inefficient at infecting human cells to being as efficient in
causing human infections as more common human influenza virus types
(Gambaryan A. et. al., 2006).
Human health risks during the H5N1 outbreak

™ Few avian influenza viruses that have crossed the species

barrier to infect humans, H5N1 has caused the largest
number of detected cases of severe disease and death in
humans.

™ In human cases associated with the ongoing H5N1

outbreaks in poultry and wild birds in Asia and parts of
Europe, the Near East and Africa, more than half of those
people reported infected with the virus have died. Most
cases have occurred in previously healthy children and
young adults and have resulted from direct or close
contact with H5N1-infected poultry or H5N1-
contaminated surfaces. In general, H5N1 remains a very
rare disease in people. The H5N1 virus does not infect
humans easily, and if a person is infected, it is very
difficult for the virus to spread to another person.

™ While there has been some human-to-human spread of

H5N1, it has been limited, inefficient and unsustained.
How flu virus invades cells and multiplies
1.The haemaglutinin (H) 2. The cell engulfs the virus to
protein on the surface of destroy it and also traps a
the flu virus binds to protease, a protein-destroying
sialic acid, a suger enzyme found in fluid outside
found on cell surface cells. The protease attacks
proteins in the virus’s haemagglutinin, but flu has
host. Most birds have a evolved to exploit this. In
different type of sialic chickens, H can usually be
acid to people, but the H activated only by a protease
on H5N1 has a mutation found in the lungs. But a
that allows it to bind to common mutation in the H
both types. allows it to be activated by a
wider variety of protease,
enabling the virus to attack all
the bird’s organs.
3. The cells pump in acid
to destroy the virus. But
again, the virus exploits
this. When the acid enters
the virus through the M2
ion channel, it triggers an
extraordinary change in
any H activated by
protease. The globular
heads of the protein fold
back and the exposed
innards bind to the cell
membrane around the
virus, making it fuse with
the viral membrane.

4. The fusion of the cell
and virus membranes
opens up a pore, and
the RNAs inside the
virus spill out into the
cell and migrate to the
cell’s nucleus.
5. Polymerase enzymes
packaged with the RNAs
churn out messenger RNA
copies of viral genes, so
the cell makes many
thousands of copies of the
10 flu proteins. Once lots
of proteins have been
made, new copies of the
viral RNAs are made.
6. New viral surface proteins –
haemaglutinin, neuraminidase
and the M2 channel- migrate
to the cell membrane as they
are produced. Here, the
neuraminidase slashes off
any sialic acids protruding
from the cell surface, so new
viruses do not stick to it and
can float free to infect other
cells.

7. The M1 matrix protein helps pack up new sets of the viral RNAs and internal
proteins and transport them to the cell membrane to join the viral surface proteins.
New viruses start budding off from the cell surface – a single infected cell can
produce 10,000 viruses.
Fig. Crystal structure of Viet04 HA and comparison with 1918 human H1, duck H5, and 1968 human H3 HAs.
(A) Overview of the Viet04 trimer, represented as a ribbon diagram. For clarity, each monomer has been
colored differently. Carbohydrates observed in the electron-density maps are colored orange, and all the
asparagines that make up a glycosylation site are labeled. Only Glu20, Glu289, and Phe154 are not labeled, as
these are on the back of the molecule. The location of the receptor binding, cleavage, and basic patch sites are
highlighted only on one monomer. (B) Structural comparison of the Viet04 monomer (olive) with duck H5
(orange) and 1918 H1 (red) HAs. Structures were first superimposed on the HA2 domain of Viet04 through the
following residues: Viet04, Gly1 to Pro160; 1918 H1 (PDB: 1rd8), Gly1 to Pro160; H3(PDB:2hmg), Gly1 to
Pro160; H5 (PDB: 1jsm ), Gly1 to Pro160. Orientation of the overlay approximates to the blue monomer in (A).
(C) Superimposition of the two long -helices of HA2 for 1918 H1 (PDB: 1rd8), avian H5 (PDB: 1jsm), human H3
(PDB: 2hmg), and Viet04 reveal that the extended interhelical loop of Viet04 is more similar to the 1918 H1 than
to the existing avian H5 structure. The side chain of Phe63 is illustrated as an example of the close proximity of
the two structures.
 Symptoms
Symptoms in Bird Symptoms in Human
¾ Change in wing color ¾ Fever,
¾ Reduction in egg ¾ Cough,
production ¾ Sore throat,
¾ Feeding problem ¾ Muscle aches,
¾ Cytokinemia ¾ Breathing problem,
¾ Pneumonia,
¾ Vomiting,
¾ Headaches etc.
 Treatment & Vaccination
y Antiviral drugs: amantadine, rimantadine, oselatmivir, and
zanamivir.

y Recombinant vaccine: sanofi-pasteur’s candidate vaccine,

Omnivest vaccine.
 Table 3.1 Confirmed human cases and mortality rate of avian influenza (H5N1)
(As of March 28, 2007)

Country Report dates Total

2003 2004 2005 2006 2007

cases deaths cases deaths cases deaths cases deaths cases deaths cases deaths

Azerbaijan 8 5 63% 8 5 63%

Cambodia 4 4 100% 2 2 100% 6 6 100%

PR China 1 1 100% 8 5 63% 13 8 62% 1 0 0% 23 14 61%

Djibouti 1 0 0% 1 0 0%

Egypt 18 10 56% 11 3 27% 29 13 45%

Indonesia 19 12 63% 56 46 82% 6 5 83% 81 63 78%

Iraq 3 2 67% 3 2 67%

Laos 2 2 100% 2 2 100%

Nigeria 1 1 100% 1 1 100%

Thailand 17 12 71% 5 2 40% 3 3 100% 25 17 68%

Turkey 12 4 33% 12 4 33%

Vietnam 3 3 100% 29 20 69% 61 19 31% 93 42 45%

Total 4 4 100% 46 32 70% 97 42 43% 116 80 69% 21 11 52% 284 169 60%

Source: World Health Organization Communicable Disease Surveillance & Response (CSR)
 Summary of the Strategic Plan
3 objectives and 5 targets
3 Objectives
• Pandemic prevention
• Reduction of morbidity and
mortality
• Development of effective respond
systems
5 Strategies and 18 measures
1. Surveillance systems
• Surveillance systems in humans and animals
• Link surveillance information of humans and
animals
• Surveillance network of human influenza
3. Pandemic Response Preparedness
• Set up standard operating procedures
• Develop the capacity of staff / volunteers
• Develop preparedness of hospitals
• Develop ca pacity of public health emergency
measures
• Develop financial measures
5. Sustainable and integrated management systems
• Pandemic alert period
• Pandemic period
5 Targets
• Surveillance system
• Pandemic response
• Stockpile of supplies and drugs
• Develop vaccine production capacity
• Public health service system
2. Stockpile of supplies
• sufficient antiviral drugs for emergency
situation
• Develop system for stockpiling and
administration of the stock
• Research and development on vaccines
and antivirals
• Develop criteria for fair distribution
4. Pubic relations and education
• Information dissemination
• Develop risk communication skills
• Set up multi sectoral working groups
• Formulate communication strategies
 Summary of the Strategic Plan
3 objectives and 5 targets

3 Objectives
• Pandemic prevention
• Reduction of morbidity and
mortality
• Development of effective respond
systems

5 Targets
• Surveillance system
• Pandemic response
• Stockpile of supplies and drugs.
• Develop vaccine production capacity
• Public health service system
 Summary of the Strategic Plan
5 Strategies and 18 measures
1.Surveillance systems
¾Surveillance systems in humans and
animals
¾Link surveillance information of humans
and animals
¾Surveillance network of human influenza
3.Pandemic Response Preparedness
¾Set up standard operating procedures
¾Develop the capacity of staff / volunteers
¾Develop preparedness of hospitals
¾Develop the capacity of public health
emergency measures
¾Develop financial measures
5.Sustainable and integrated management systems
¾Pandemic alert period
¾Pandemic period
2. Stockpile of supplies and drugs
¾Sufficient antiviral drugs for emergency
situation
¾Develop system for stockpiling and
administration of the stock
¾Research development on vaccines
and antiviral drug
¾Develop criteria for fair distribution
4.Pubic relations and education
¾Develop risk communication skills
¾Information dissemination
¾Set up multi sectoral working groups
¾Formulate communication strategies
Roles & responsibility of concerned agencies

¾ Ministry of public health

¾ Department of livestock development
¾ Ministry of defense
¾ Department of disaster prevention and
mitigation
¾ National police bureau
¾ Public relation department
¾ Ministry of education
 Conclusion
Avian influenza viruses do not usually infect humans;
however, several instances of human infections and
outbreaks of avian influenza have been reported since 1997.
In 2003, influenza A (H7N7) infections occurred in the
Netherlands among persons, more than 80 cases of H7N7
illness were confirmed by testing and one patient died. It is
believed that most cases of avian influenza infection in
humans have resulted from contact with infected poultry or
contaminated surfaces. When highly pathogenic influenza H5
or H7 viruses cause outbreaks, between 90% and 100% of
poultry can die from infection and then quarantine and
depopulation (or culling) and surveillance around affected
flocks is the preferred control and eradication option.
The following interim recommendations are based on what
are deemed optimal precautions for protecting individuals
involved in the care of patients with highly pathogenic
avian influenza from illness and for reducing the risk of viral
reassortment (i.e., mixing of genes from human and avian
viruses).

i. Standard Precautions
Pay careful attention to hand hygiene before and after all
patient contact or contact with items potentially
contaminated with respiratory secretions.

ii. Contact Precautions

Use gloves and gown for all patient contact, also Use
dedicated equipment such as stethoscopes,
disposable blood pressure cuffs, disposable thermometers,
etc.
iii. Eye protection
Wear goggles or face shields when within 3 feet of the
patient.

iv. Airborne Precautions

Place the patient in an airborne isolation room (AIR).

v. Vaccination of Health-Care Workers against Human

Influenza

vi. Surveillance and Monitoring of Health-Care Workers

Instruct health-care workers to be vigilant for the
development of fever, respiratory symptoms, and/or
conjunctivitis (i.e., eye infections) for 1 week after last
exposure to avian influenza-infected patients.
THANKS TO ALL