ADENOVIRUS GENOME AND ITS REPLICATION

Adenovirus Genome
Adenoviruses are nonenveloped, linear double stranded DNA viruses. Its genome is completely surrounded by an icosohedral protein capsid build up of three major proteins group which are hexon, penton base and knobbed fiber.

The length of adenovirus linear genome is between 26 and 45 Kbp and therefore allows the virus to theoretically carry 22 to 40 genes. However, bigger genome size does not change the fact that this virus is still an obligate intracellular parasite that fully depends on the host cell machinery to survive and replicate.

Generally, this linear virus genome is about 36 Kbp in size and consists of immediate early (E1A), early (E1-E4), intermediate and late genes (L1-L5) (Figure 1). Transcription of these genes can be divided into early and late phase, respectively, occurring before or after DNA replication.




Figure 1 Adenovirus genome

An interesting and yet advantageous feature of this viral genome is that it has a terminal 55 kDa protein associated with each of the 5' ends of the linear dsDNA. Such protein functions as primers in viral replication and ensures that the ends of the virus' linear genome are adequately replicated.

Adenovirus Replication
The replication strategy of the virus depends on the nature of its genome, since different groups of viruses classify under Baltimore system will undergo different replication cycles due to the nature of their genome. Adenovirus is classified as Class I dsDNA virus with the following replication cycle.



Replication Cycle of Adenovirus

1. Attachment
Uptake process of the virion particle by the host cell. Two stage process involving an initial interaction of the fibre protein with a range of cellular receptors, which include the MHC class I molecule and the coxsackievirus-adenovirus receptor. The penton base protein then binds to the integrin family of cell surface heterodimers allowing internalization via receptor-mediated endocytosis. Most cells express primary receptors for the adenovirus fibre coat protein, however internalisation is more selective.

2. Penetration

Phagocytosis of the whole adenovirus virion into host cell cytoplasm as a phagocytic vacuole. The toxic activity of the pentons is responsible for rupture of the phagocytic membrane and release the virion particle into the cytoplasm.


3. Uncoating

Follows an ordered sequence, first the pentons and capsids are digested by the host cell lysozyme, releasing a spherical, partially uncoated particle into the cytoplasm. The core migrates to the nucleus through nuclear pores with the help of cellular microtubules, whereupon it is converted into a virus DNA-cell histone complex.



Figure 2 Attachment, penetration and uncoating steps in adenovirus replication

Gene Expression

Prior to DNA replication, immediate early and early mRNAs are transcribed from the input DNA. Transcription of the Adenovirus genome is regulated by virus-encoded trans-acting regulatory factors (products of the immediate early genes regulate expression of the early genes).
Phase Gene transcribed Protein translated
Immediate early, E1A trans-acting transcriptional regulatory factor
Early E1B, E2A, E2B, E3, E4, Regulatory factors and some virion proteins
Late Late genes , mostly virion proteins eg capsid, pentons etc




Figure 3 Gene expression of adenovirus genome

DNA Replication
The steps involved in adenovirus DNA replication can be summarized as follows:
• First, the viral genome is coated with DBP.
• This protein reacts co-operatively with the cellular transcription factor, NFI which binds to a recognition site within the origin of replication, separated from the 1-18 bp core by a precisely defined spacer region.
• NFIII also binds at a specific recognition site between nucleotides 39 and 48.
• Protein-protein interactions, between NFI and pol, and pTP and NFIII help recruit the pTP-pol heterodimer into the preinitiation complex.
• Interaction between the heterodimer and specific base pairs 9 to 18 in the DNA sequence ensures correct positioning and the complex is further stabilized by interactions between the incoming pTP-pol and the genome-bound TP.
• DNA replication is then initiated by a protein priming mechanism in which a covalent linkage is formed between the alpha-phosphoryl group of the terminal residue, dCMP and the beta-hydroxyl group of a serine residue in pTP, a reaction catalysed by pol. This acts as a primer for synthesis of the nascent strand.
• Base pairing with the second GTA triplet of the template strand guides the synthesis of a pTP-trinucleotide, which then jumps back 3 bases, to base pair with the first triplet (also GTA) and synthesis then proceeds by displacing the non-template strand.
• NFI dissociates as the first nucleotide binds just prior to the initiation reaction. Dissociation of pTP from pol begins as the pTP-trinucleotide is formed and is almost complete by the time 7 nucleotides have been synthesized.
• NFIII dissociates as the replication binding fork passes through the NFIII binding site.



Figure 4 Adenovirus DNA Replication



Assembly of Virion Particles and Lysis of Host Cell
The late phase of the adenovirus life cycle is focused on producing sufficient quantities of structural protein to pack all the genetic material produced by DNA replication. Once the viral components have successfully been replicated the virus is assembled into its protein shells and released from the cell as a result of virally induced cell lysis.



Figure 5 Summary of adenovirus replication cycle

TREATMENT AND PREVENTION OF ADENOVIRUS

Up to date, there is no vaccine against adenovirus infections. The treatments of adenovirus infection are supportive and aim on relieving the symptoms that related with the infection.

Diagnostic
Laboratory diagnostic for adenovirus can be generally divides into three types as below:


1. Direct detection of adenovirus:
Virus particle of adenovirus can be detected by observing under electrotron microscope of fecal extract. Specific immunochemistry method such as ELISA also can be used to detect adenovirus antigen. Last but not least, adenovirus nucleic acid can detect by Polymerase chain reaction using tissue sample and body fluid.

2. Isolation
Isolation of adenovirus can be from various sources such as throat, conjunctival swab and urines. Other sources of sample such as stool and rectal swab also can be used but is more difficult to isolate the adenovirus.

3. Serology
Serology test involve use of blood to detect infection of adenovirus using haemaglutination inhibition. Detection using specific antibodies following infection namely neutralization test. Both of method state as above is known as adenovirus typing method.




Treatment

The infection of adenovirus cannot be treated with antibiotic and no need to have therapy. What we can do is treating symptoms and complications of the infection. Via this treatment strategy, death is rare in adenovirus infection. It is difficult to differentiate from similar disease caused by a variety of RNA viruses.

Before 1996, there are adenovirus vaccines that are safe and effective for serotype 4 and 7. These vaccines are only available for preventing ARD among US military. However, this vaccine was stop in 1996. Antiviral has not been tried to produce for the adenovirus-induced respiratory syndromes.

Treatment for respiratory infection caused by adenovirus may include:
• Increase fluid intake into patient’s body. Drink a lot of water or if necessary, an intravenous (IV) line and electrolyte may be used.
• Bronchodilator medications through an inhaler can be used to open your airways.
• Mechanical ventilation such as oxygen through a mask, nasal progs or an oxygen tent to assist patient’s breathing.

Treatment for intestinal infection may include:
• Oral rehydration by consuming water, formula milk, electrolyte-containing fluids, soda, juice and others.
• Intravenous fluid and nasogatic (NG) tube may be used. The electrolyte level can be monitored.


Prevention of adenovirus

In adenovirus infection, the key prevention is personal hygiene. Careful hand washing can keep away from adenovirus infection. Disinfection of environmental surface with Clorox also keeps clean the adenovirus. Conjunctivitis in swimming pool can be prevented by addition of adequate level of chlorine in the water.

INFECTIONS OF ADENOVIRUSES.

The infections of adenoviruses into the cells involve specific interactions between the receptor of the cell surface and viral capsid protein. Adenoviruses infect most of the cells by attachment of the knob that projected from the capsid of the viruses to the specific receptor on the surface of the cell membrane. Naturally, receptors that responsible in the attachment of the adenoviruses are known as coxsackie and adenovirus receptor (CAR), but there are also certain serotypes that attach to other types of receptor.
To date, there are 49 distinct types of adenoviruses have been linked to human infection and more than 120 adenoviruses are known to infect a wide variety of vertebrate species that include mammals, fish, birds, amphibians and reptiles. Each of these viruses produces unique immune responses. Therefore, if an individual develops immunity to one type of virus after an infection, he or she can still become infected with one of the other types. Adenoviruses are unusually stable and can survive for prolonged periods outside the body.
Human adenoviruses serotypes are classified into six distinct species which designated from A through F. Human adenoviruses are mostly involved in mild and self-limiting disease in immunocompetent individuals, however the disease may be acute or even life-threatening in children an immunocompromised adults. Generally, different species of human adenoviruses will exhibit distinct tissue tropism and clinical manifestation.
Most of the infections that involved adenoviruses will give major causes in respiratory tract infections. Normally, the species (subgroup) that lead to this infection are adenoviruses from the subgroup of C, E and B1. Besides that, those of subgroup D and E of this adenovirus may also lead to the infections in ocular parts. Human adenovirus serotypes from subgroup A and F are reported to be a major responsible for the infections of gastrointestinal whereas the subgroups of B2 of the adenovirus will cause in renal and urinary tract infections but it is rarely happened.


Adenoviral infections affect infants and young children much more frequently than adults. Some children (especially the small ones) can develop adenovirus bronchiolitis or pneumonia, both of which can be severe. Adenoviruses can also cause coughing fits that look almost exactly like whooping cough especially in babies. Most of the infections by adenoviruses in human are recovered by themselves, but some people that suffered from the immunodeficiency sometimes die due to this adenovirus infections, and rarely even previously healthy people can die from these infections.
The majority of the population will have experienced at least one adenoviral infections by age 10. Most adenoviral infections in kid are take place in the first year of life although this infection can occur at any age. There are possibilities that repeated infections occurred due to many different types and species of adenovirus.
Other illnesses that associated with adenovirus include:
• encephalitis (inflammation of the brain) and other infections of the central nervous system (CNS)
• gastroenteritis (inflammation of the stomach and intestines), which sometimes leads to enlarged lymph nodes in the intestines and rarely intussusception
• acute pharyngoconjunctival fever (inflammation of the lining of the eye [conjunctivitis] with fever)
• acute mesenteric lymphadenitis (inflammation of lymph glands in the abdomen)
• chronic interstitial fibrosis (abnormal growth of connective tissue between cells)
• intussusception (a type of intestinal obstruction)
• pneumonia that does not respond to antibiotic therapy
• whooping cough syndrome when Bordetella pertussis (the bacterium that causes classic whooping cough) is not found

adenovirus tropism and receptor usage.

ADENOVIRUS TRANSMISSION

Adenovirus Transmission

Adenoviruses are very common viruses that can cause infections. There are over fifty subtypes of adenovirus. They most commonly cause upper respiratory tract infections, including the common cold, sore throats, tonsillitis, ear infections, and conjunctivitis. Another common adenoviral infection is pharyngoconjunctival fever (sore throat, red eyes, and a fever).
Virus transmission is the passing of a disease from an infected individual or group to a previously uninfected individual or group. The virus that causes disease may be transmitted from one person to another. Adenovirus can spread via direct contact, droplet transmission, respiratory droplet and fecal-oral transmission. Basically the adenovirus can be transmitted by:

1. Ingestion / oral fecal route
2. Respiration through respiratory route
3. Contact/Hand-to-eye transfer
4. Venereal

Ingestion / oral fecal route

Fecal–oral transmission occurs when the virions found in the stool of one person (or animal) are swallowed by another person. Transmission can spread directly through person that handling utensils that been contaminate with the virions or even contaminated food and beverages. Fecal material can be ingested through contamination of water supplies, poor hand washing between the bathroom and the kitchen, eating food contaminated by houseflies, or poor hygiene after handling diapers. Swimming pools and water parks can also be locations of fecal–oral transmission. If the water is not visibly contaminated and is adequately chlorinated, just getting the water in the mouth is usually enough to cause transmission of adenovirus; the risk is greatly increased by swallowing.

Respiratory droplets and body fluids

Adenoviruses are usually stable to chemical or physical agents and adverse pH conditions, allowing for prolonged survival outside of the body and water. Droplet transmission occurs when thee viruses travel on relatively large respiratory droplets that people sneeze, cough, drip, or exhale. These droplets are loaded with infectious particles.

They can be spread directly if people are close enough to each other. More often, though, fomites are involved. The droplets land on hands, toys, tables, mats, or other surfaces, where they sometimes remain infectious for hours. Hands that come in contact with these surfaces (doorknobs, telephones, pens, etc.) become contagious. When the infectious hand touches the nose or eyes, the infection is able to enter the new person.

Contact/Hand-to-eye transfer

Adenovirus is highly contagious, as indicated by the occurrence of multiple cases in situations of close contact, such as schools, hospitals, community centres and food court. The type of adenovirus that cause respiratory and intestinal infection basically spread from person to person through respiratory secretions (coughs or sneezes) or fecal contamination.

A person might also get the virus by holding hands or sharing utensils with an infected person. Indirect transmission can occur through exposure to the contaminated surfaces of furniture and other objects. The types of adenovirus causing conjunctivitis (acute inflammation of the conjunctiva) may be transmitted by sharing contaminated objects or by touch.

Sexual Transmission

This type of transmission occurs during sexual activity with another person. Transmission is either directly between surfaces in contact during intercourse (the usual route for bacterial infections and those infections causing sores) or from secretions such as semen or fluid which carry infectious agents that get into the partner's blood stream and leading to the virions transmission into the person body and starting the disease symptoms.

CLASSIFICATION, DIVERSITY AND STRUCTURE OF ADENOVIRUS

CLASSIFICATION

Adenovirus can be classified into fives genera that are Genus Atadenovirus, Genus Aviadenovirus, Genus Ichtadenovirus, Genus Mastadenovirus and Genus Siadenovirus.

Genus Atadenovirus

Atadenoviruses infect animal with a broad host range from several vertebrate classes. It has capsid fibers, a core, and associated protein. Their capsids are not enveloped but have icosahedral symmetry. The isometric capsid has a diameter of 80-100 nm. The capsid shells of virions are composed of a single layer. Capsids appear hexagonal in outline.

Genus Aviadenovirus

These types of virus are mainly infected bird species.

Genus Ichtadenovirus

This family of adenovirus share very similar characteristic with other genus such as the genome has terminally redundant sequences which have inverted terminal repetitions (ITR).its structure also very similar to Aviandenovirus. The terminal protein (TP) is covalently attached to each end of the genome. But, their hosts are not very known for now.

Genus Mastadenovirus

This virus can cause respiratory infections and acute follicular conjunctivitis to many species including human.

Genus Siadenovirus

This virus comprising species such as frog and turkey.

The genome of all this genus share same similar characteristic that are their genomes are non segmented, linear double-stranded DNA of 35-36kb encoding about 40 proteins. The genome has terminally redundant sequences which have inverted terminal repetitions (ITR). The terminal protein (TP) is covalently attached to each end of the genome.

DIVERSITY

Diversity of adenovirus is very complex. There are 54 types of human adenovirus comprising of seven species (A to G).
• A: 12, 18, 31
• B: 3, 7, 11, 14, 16, 34, 35, 50
• C: 1, 2, 5, 6
• D: 8, 9, 10, 13, 15, 17, 19, 20, 22, 23, 24, 25, 26, 27, 28, 29, 30, 32, 33, 36, 37, 38, 39, 42, 43, 44, 45, 46, 47, 48, 49, 51, 53, 54
• E: 4
• F: 40, 41
• G: 52

Adenovirus serotype 14 in types B is known to cause potentially fatal adenovirus infections. This type of virus can cause severe respiratory infection.
For adenovirus serotype 36, it is also known to infect human being. It was first isolates from the girl that have diabetes and enteritis. It is also cause respiratory and eye infection to human. The relationship between body fat and this type of virus have a positive correlation.
Other types of adenovirus can cause several diseases such as species B and C can cause respiratory disease to human. Species B and D also can cause conjunctivitis to human and serotype 4o and 41 in species F can cause gastroenteritis.



STRUCTURE

The structure of adenovirus genome are Non segmented, linear double-stranded DNA of 35-36kb. The genome has terminally redundant sequences which have inverted terminal repetitions (ITR). The terminal protein (TP) is covalently attached to each end of the genome.






This virus also contained capsid as their genome protection. Adenoviruses are double-stranded DNA viruses. They have icosahedral capsids with twelve vertices and seven surface proteins. The virion is non-enveloped, spherical and about seventy to ninety nm in size. The genome encodes about thirty proteins. This virus also consisted of spike for infection.