Influenza H1N1 A: A close insight

Dr. Mustafa AbabnehMolecular Virologist

Outlines

• Virus structure, proteins and functions• Influenza A replication• Influenza A Reassortment• Identification of reassortment• Alignment of Current H1N1 influenza viruses ,

and between H1N1 and H5N1.• Mutational effects

Virus charateristics• Orthomyxoviridae Family• Has a segmented negative-sense, single-stranded RNA viruses• total genome length is 12000-15000 nt.• 16 H antigens (H1 - H16) and 9 N antigens (N1 - N9), which can be found

in all possible combinations. • The nucleoprotein antigen (A, B, C) determines the virus type. The HA

and NA antigens determine subtypes.Influenza virus A:both antigen shift and antigen drift noted. High antigenic variability in the

surface glycoproteins HA and NA. Has 8 segments.

Influenza virus B : Members infect only humans; associated with epidemics; antigenic drift noted. Has 8 segments

Influenza virus C : human and swine, Has 7 segments.

Replication of orthomyxovirus

• Replication starts with the binding of HA protein to the sialic acid receptors

• Then penetration and uncoating• Genome Replication takes place in the nucleus• Cap-snatching• NOT in the cytoplasm

Antigenic variation

• Antigenic shift:• These are the major changes based on the

reassortment of segments of the genome. In the reassortment entire segments of RNA can be exchanged between two or more viruses infecting the same host cell.

• Antigenic drifts: these are minor changes caused by point mutation in the genes encoding HA or NA glycoproteins

• Antigenic shift will render the resulting virus the ability to cross species barriers.

• Antigenic drift will render the virus to transmit efficiently between the same species. RNA dependent RNA polymerase

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Influenza Reassortment

Reassortment in H1N1 influenza ASegment Size in amino acid Species origin Geographical origin

PB2: polymerase basic subunit 2

760 a.a Avian North America

PB1 :polymerase basic subunit 1

758 a.a Human 1993:H3N2 Human

PA :polymerase acid

717 a.a Avian North America

HA : Heamagglutinin

567 a.a Swine (H1) North America

NP: nucleoprotein 499 a.a Swine North America

NA: neurominidase 470 a.a Swine (N1) Europe

M : matrix (M1,M2) 324 a.a Swine Euroasia

NS : Non-structral proteins (NS1,NS2)

279 a.a swine North America

How we can Know the reassortments events

Multiplex RT-PCR identification of reassortment

Influenza A

• Current circulating strains are H1N1 and H3N2• Human subtypes include H1N1, H3N2, H1N2,

and H2N2• Avian subtypes include H1 to H16 and N1 to

N9• Bird human H5N1, H9N2, H7N7, H7N2,

H7N3

H1N1 sequence database

H1N1 genes sequences

PB1 (H1N1)

PB2 PA HA NP NA M NS

(H5N1)Jordan-2006

95.9% 84.5% 96.1% 9.7% 93.6% 87.6% 10.1% 77.4%

Sequence similarity between current H1N1 and H5N1 (Jordan-2006)

• Alignment was done with ClustalW, lasergene DNA analysis Software.

Sequence similarity between current H1N1 isolates

H1N1-Cal PB2 PB1 PA HA NP NA M NS

Texas 99.7% 99.9% 99.9% 99.8% 100% 99.9% 99.9% 99.9%

Denmark N.A N.A N.A 99.5% N.A 99.9% 100% 100%

Auckland N.A N.A N.A N.A N.A 99.6% 100% N.A

Germany N.A N.A N.A N.A N.A 99.9% N.A N.A

Mutational analysis• Amino acid markers of host specificity ,Does

Influenza H1N1 A has the markers?• Tamiflu resistant Mutation• NS1 mutation- glutamic acid in position 92.

• by compare the amino acids sequences of all influenza pandemic strains.

• If positions: 87 NS1, 103 NS1, 207 NS1 and 63 NS2 contains Human Amino acid: this means the virus will be cross to human from other species.

Tamiflu resistant H1N1

• H275Y • Histidine to tyrosine subsiotiution in the

neuraminidase active site (H 275Y in N1 )• Detection by sequencing.• The current H1N1 still Tamiflu sensitive

Tamiflu reseistant H1N1

Tamiflu resistant mutation in current H1N1

Tamiflu resistant mutation in Human H1N1 (Human strain)

NS1 H5N1 (D 92 E)

NS1 H5N1 (D 92 E)

NS1 of H1N1 (D 92 E)

Refernces

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