Phylogenetic analysis of influenza A viruses (H3N2) circulating in Zhytomyr region during 2013–2014 epidemic season
Aim. To perform phylogenetic analysis of the hemagglutinin (HA) and neuraminidase (NA) genes of influenza A(H3N2) viruses circulating in the Zhytomyr region during 2013–2014 epidemic season. To make comparison of the HA and NA genes sequences of the Zhytomyr region isolates with the HA and NA genes...
Saved in:
| Published in: | Вiopolymers and Cell |
|---|---|
| Date: | 2015 |
| Main Authors: | , , , , , |
| Format: | Article |
| Language: | English |
| Published: |
Інститут молекулярної біології і генетики НАН України
2015
|
| Subjects: | |
| Online Access: | https://nasplib.isofts.kiev.ua/handle/123456789/152475 |
| Tags: |
Add Tag
No Tags, Be the first to tag this record!
|
| Journal Title: | Digital Library of Periodicals of National Academy of Sciences of Ukraine |
| Cite this: | Phylogenetic analysis of influenza A viruses (H3N2) circulating in Zhytomyr region during 2013–2014 epidemic season / O.G. Boyalska, I.M. Kyrychuk, I.G. Budzanivska, A.P. Mironenko, L.V. Radchenko, O.Yu. Smutko // Вiopolymers and Cell. — 2015. — Т. 31, № 3. — С. 226-232. — Бібліогр.: 15 назв. — англ. |
Institution
Digital Library of Periodicals of National Academy of Sciences of Ukraine| id |
nasplib_isofts_kiev_ua-123456789-152475 |
|---|---|
| record_format |
dspace |
| spelling |
Boyalska, O.G. Kyrychuk, I.M. Budzanivska, I.G. Mironenko, A.P. Radchenko, L.V. Smutko, O.Yu. 2019-06-11T18:55:45Z 2019-06-11T18:55:45Z 2015 Phylogenetic analysis of influenza A viruses (H3N2) circulating in Zhytomyr region during 2013–2014 epidemic season / O.G. Boyalska, I.M. Kyrychuk, I.G. Budzanivska, A.P. Mironenko, L.V. Radchenko, O.Yu. Smutko // Вiopolymers and Cell. — 2015. — Т. 31, № 3. — С. 226-232. — Бібліогр.: 15 назв. — англ. 0233-7657 DOI: http://dx.doi.org/10.7124/bc.0008E4 https://nasplib.isofts.kiev.ua/handle/123456789/152475 575.22 + 578.53 Aim. To perform phylogenetic analysis of the hemagglutinin (HA) and neuraminidase (NA) genes of influenza A(H3N2) viruses circulating in the Zhytomyr region during 2013–2014 epidemic season. To make comparison of the HA and NA genes sequences of the Zhytomyr region isolates with the HA and NA genes sequences of influenza viruses circulating in the world. Methods. Laboratory diagnosis was conducted by real-time polymerase chain reaction (RT-PCR). In this study the sequencing and phylogenetic analysis were carried out. Results. For the first time the genes of influenza A(H3N2) viruses isolated in the Zhytomyr region during 2013–2014 epidemic season, coding hemagglutinin and neuraminidase were compared with their orthologs. According to the results of this comparison the phylogenetic tree was constructed. Additionally, the amino acid substitutions of the influenza viruses circulating in Ukraine and worldwide were analyzed. Conclusions. The nucleotide sequences of the influenza A(H3N2) viruses genes HA and NA isolated in the Zhytomyr region were identified. Based on the nucleotide sequences of HA and NA we constructed the influenza virus phylogenetic tree demonstrating that the virus isolated in the Zhytomyr region was closely related to the Ukrainian isolate from Kharkov and in the world to the isolates from Germany, Romania, Italy. Мета. Зробити філогенетичний аналіз генів гемаглютиніну (HA) і нейрамінідази (NA) вірусів грипу А(H3N2), що були поширені в Житомирській області під час епідемічного сезону 2013–2014рр. і порівняти їх з тими, що циркулювали в світі. Методи. Лабораторну діагностику здійснювали за допомогою ПЛР у реальному часі (real-time RT-PCR). Було проведено секвенування та філогенетичний аналіз. Результати. В епідемічному сезоні 2013–2014 рр. вперше було зроблено секвенування генів НА та NA вірусів грипу А(H3N2), виділених з клінічного матеріалу хворих з Житомирської області та в подальшому увійшли до бази даних вірусів грипу з усього світу (GISAID). Висновки. Житомирські ізоляти розташувались поряд з українським ізолятом з Харкова, а серед світових поряд з ізолятами з Німеччини, Румунії, Італії. В послідовностях генів гемаглютиніну було виявлено заміщення I214T. Виділені нами ізоляти в епідемічному сезоні 2013–2014рр. мали високу генетичну спорідненість (99 %) до вірусів, що знаходяться в групі на філогенетичному дереві, тому що мають синонімічне заміщення. Житомирські ізоляти розташувались у домінуючій групі в світі 3С групи 3 генетичного кластеру A/Victoria/208 і були подібними до вакцинного штаму A/Texas/ 50/2012. В послідовностях генів нейрамінідази суттєвих заміщень виявлено не було. Житомирські ізоляти, як і багато українських та світових ізолятів виявились подібними до референс-штаму A/AthensGR/112/2012, ніж до більш нових референс-вірусів. Цель. Провести филогенетический анализ генов гемагглютинина (HA) и нейраминидазы (NA) вирусов гриппа А(H3N2), которые циркулировали в Житомирской области во время эпидемического сезона 2013–2014гг. и сравнить их с таковыми со всего мира. Методы. Лабораторную диагностику проводили с помощью ПЦР в реальном времени (real-time RT-PCR). Было проведено секвенирование и филогенетический анализ. Результаты. В эпидемическом сезоне 2013–2014гг. впервые было сделано секвинирование генов НА та NA вирусов гриппа А(H3N2), виделенных из клинического материала больных в Житомирськой области и в дальнейшем вошли в базу данных вирусов гриппа со всего мира (GISAID). Выводы. Житомирские изоляты разместились рядом с украинским изолятом из Харькова, а среди мировых рядом с изолятами из Германии, Румынии, Италии. В последовательностях генов гемагглютинина было выявлено замещение I214T. Выделенные нами изоляты в эпидемическом сезоне 2013–2014гг. имели высокое генетическое сходство (99 %) к вирусам, которые находяться в группе на филогенетическом дереве, так как имеют синонимическое замещение. Житомирские изоляты разместились в доминирующей субгруппе в мире 3С группы 3 генетического кластера A/ Victoria/208. В последовательностях генов нейраминидазы существенных мутаций не выявлено. Житомирские изоляты, как много украинских и мирових изолятов оказались подобными к референс-штамму A/AthensGR/112/2012, чем к более новым референс-вирусам. en Інститут молекулярної біології і генетики НАН України Вiopolymers and Cell Viruses and Cell Phylogenetic analysis of influenza A viruses (H3N2) circulating in Zhytomyr region during 2013–2014 epidemic season Філогенетичний аналіз вірусів грипу А(H3N2), що циркулювали в Житомирській області протягом епідемічного сезону 2013–2014рр. Филогенетический анализ вирусов гриппа А(H3N2) которые циркулировали в Житомирской области в период эпидемического сезона 2013–2014гг. Article published earlier |
| institution |
Digital Library of Periodicals of National Academy of Sciences of Ukraine |
| collection |
DSpace DC |
| title |
Phylogenetic analysis of influenza A viruses (H3N2) circulating in Zhytomyr region during 2013–2014 epidemic season |
| spellingShingle |
Phylogenetic analysis of influenza A viruses (H3N2) circulating in Zhytomyr region during 2013–2014 epidemic season Boyalska, O.G. Kyrychuk, I.M. Budzanivska, I.G. Mironenko, A.P. Radchenko, L.V. Smutko, O.Yu. Viruses and Cell |
| title_short |
Phylogenetic analysis of influenza A viruses (H3N2) circulating in Zhytomyr region during 2013–2014 epidemic season |
| title_full |
Phylogenetic analysis of influenza A viruses (H3N2) circulating in Zhytomyr region during 2013–2014 epidemic season |
| title_fullStr |
Phylogenetic analysis of influenza A viruses (H3N2) circulating in Zhytomyr region during 2013–2014 epidemic season |
| title_full_unstemmed |
Phylogenetic analysis of influenza A viruses (H3N2) circulating in Zhytomyr region during 2013–2014 epidemic season |
| title_sort |
phylogenetic analysis of influenza a viruses (h3n2) circulating in zhytomyr region during 2013–2014 epidemic season |
| author |
Boyalska, O.G. Kyrychuk, I.M. Budzanivska, I.G. Mironenko, A.P. Radchenko, L.V. Smutko, O.Yu. |
| author_facet |
Boyalska, O.G. Kyrychuk, I.M. Budzanivska, I.G. Mironenko, A.P. Radchenko, L.V. Smutko, O.Yu. |
| topic |
Viruses and Cell |
| topic_facet |
Viruses and Cell |
| publishDate |
2015 |
| language |
English |
| container_title |
Вiopolymers and Cell |
| publisher |
Інститут молекулярної біології і генетики НАН України |
| format |
Article |
| title_alt |
Філогенетичний аналіз вірусів грипу А(H3N2), що циркулювали в Житомирській області протягом епідемічного сезону 2013–2014рр. Филогенетический анализ вирусов гриппа А(H3N2) которые циркулировали в Житомирской области в период эпидемического сезона 2013–2014гг. |
| description |
Aim. To perform phylogenetic analysis of the hemagglutinin (HA) and neuraminidase (NA) genes of influenza A(H3N2) viruses circulating in the Zhytomyr region during 2013–2014 epidemic season. To make comparison of the HA and NA genes sequences of the Zhytomyr region isolates with the HA and NA genes sequences of influenza viruses circulating in the world. Methods. Laboratory diagnosis was conducted by real-time polymerase chain reaction (RT-PCR). In this study the sequencing and phylogenetic analysis were carried out. Results. For the first time the genes of influenza A(H3N2) viruses isolated in the Zhytomyr region during 2013–2014 epidemic season, coding hemagglutinin and neuraminidase were compared with their orthologs. According to the results of this comparison the phylogenetic tree was constructed. Additionally, the amino acid substitutions of the influenza viruses circulating in Ukraine and worldwide were analyzed. Conclusions. The nucleotide sequences of the influenza A(H3N2) viruses genes HA and NA isolated in the Zhytomyr region were identified. Based on the nucleotide sequences of HA and NA we constructed the influenza virus phylogenetic tree demonstrating that the virus isolated in the Zhytomyr region was closely related to the Ukrainian isolate from Kharkov and in the world to the isolates from Germany, Romania, Italy.
Мета. Зробити філогенетичний аналіз генів гемаглютиніну (HA) і нейрамінідази (NA) вірусів грипу А(H3N2), що були поширені в Житомирській області під час епідемічного сезону 2013–2014рр. і порівняти їх з тими, що циркулювали в світі. Методи. Лабораторну діагностику здійснювали за допомогою ПЛР у реальному часі (real-time RT-PCR). Було проведено секвенування та філогенетичний аналіз. Результати. В епідемічному сезоні 2013–2014 рр. вперше було зроблено секвенування генів НА та NA вірусів грипу А(H3N2), виділених з клінічного матеріалу хворих з Житомирської області та в подальшому увійшли до бази даних вірусів грипу з усього світу (GISAID). Висновки. Житомирські ізоляти розташувались поряд з українським ізолятом з Харкова, а серед світових поряд з ізолятами з Німеччини, Румунії, Італії. В послідовностях генів гемаглютиніну було виявлено заміщення I214T. Виділені нами ізоляти в епідемічному сезоні 2013–2014рр. мали високу генетичну спорідненість (99 %) до вірусів, що знаходяться в групі на філогенетичному дереві, тому що мають синонімічне заміщення. Житомирські ізоляти розташувались у домінуючій групі в світі 3С групи 3 генетичного кластеру A/Victoria/208 і були подібними до вакцинного штаму A/Texas/ 50/2012. В послідовностях генів нейрамінідази суттєвих заміщень виявлено не було. Житомирські ізоляти, як і багато українських та світових ізолятів виявились подібними до референс-штаму A/AthensGR/112/2012, ніж до більш нових референс-вірусів.
Цель. Провести филогенетический анализ генов гемагглютинина (HA) и нейраминидазы (NA) вирусов гриппа А(H3N2), которые циркулировали в Житомирской области во время эпидемического сезона 2013–2014гг. и сравнить их с таковыми со всего мира. Методы. Лабораторную диагностику проводили с помощью ПЦР в реальном времени (real-time RT-PCR). Было проведено секвенирование и филогенетический анализ. Результаты. В эпидемическом сезоне 2013–2014гг. впервые было сделано секвинирование генов НА та NA вирусов гриппа А(H3N2), виделенных из клинического материала больных в Житомирськой области и в дальнейшем вошли в базу данных вирусов гриппа со всего мира (GISAID). Выводы. Житомирские изоляты разместились рядом с украинским изолятом из Харькова, а среди мировых рядом с изолятами из Германии, Румынии, Италии. В последовательностях генов гемагглютинина было выявлено замещение I214T. Выделенные нами изоляты в эпидемическом сезоне 2013–2014гг. имели высокое генетическое сходство (99 %) к вирусам, которые находяться в группе на филогенетическом дереве, так как имеют синонимическое замещение. Житомирские изоляты разместились в доминирующей субгруппе в мире 3С группы 3 генетического кластера A/ Victoria/208. В последовательностях генов нейраминидазы существенных мутаций не выявлено. Житомирские изоляты, как много украинских и мирових изолятов оказались подобными к референс-штамму A/AthensGR/112/2012, чем к более новым референс-вирусам.
|
| issn |
0233-7657 |
| url |
https://nasplib.isofts.kiev.ua/handle/123456789/152475 |
| citation_txt |
Phylogenetic analysis of influenza A viruses (H3N2) circulating in Zhytomyr region during 2013–2014 epidemic season / O.G. Boyalska, I.M. Kyrychuk, I.G. Budzanivska, A.P. Mironenko, L.V. Radchenko, O.Yu. Smutko // Вiopolymers and Cell. — 2015. — Т. 31, № 3. — С. 226-232. — Бібліогр.: 15 назв. — англ. |
| work_keys_str_mv |
AT boyalskaog phylogeneticanalysisofinfluenzaavirusesh3n2circulatinginzhytomyrregionduring20132014epidemicseason AT kyrychukim phylogeneticanalysisofinfluenzaavirusesh3n2circulatinginzhytomyrregionduring20132014epidemicseason AT budzanivskaig phylogeneticanalysisofinfluenzaavirusesh3n2circulatinginzhytomyrregionduring20132014epidemicseason AT mironenkoap phylogeneticanalysisofinfluenzaavirusesh3n2circulatinginzhytomyrregionduring20132014epidemicseason AT radchenkolv phylogeneticanalysisofinfluenzaavirusesh3n2circulatinginzhytomyrregionduring20132014epidemicseason AT smutkooyu phylogeneticanalysisofinfluenzaavirusesh3n2circulatinginzhytomyrregionduring20132014epidemicseason AT boyalskaog fílogenetičniianalízvírusívgripuah3n2ŝocirkulûvalivžitomirsʹkíioblastíprotâgomepídemíčnogosezonu20132014rr AT kyrychukim fílogenetičniianalízvírusívgripuah3n2ŝocirkulûvalivžitomirsʹkíioblastíprotâgomepídemíčnogosezonu20132014rr AT budzanivskaig fílogenetičniianalízvírusívgripuah3n2ŝocirkulûvalivžitomirsʹkíioblastíprotâgomepídemíčnogosezonu20132014rr AT mironenkoap fílogenetičniianalízvírusívgripuah3n2ŝocirkulûvalivžitomirsʹkíioblastíprotâgomepídemíčnogosezonu20132014rr AT radchenkolv fílogenetičniianalízvírusívgripuah3n2ŝocirkulûvalivžitomirsʹkíioblastíprotâgomepídemíčnogosezonu20132014rr AT smutkooyu fílogenetičniianalízvírusívgripuah3n2ŝocirkulûvalivžitomirsʹkíioblastíprotâgomepídemíčnogosezonu20132014rr AT boyalskaog filogenetičeskiianalizvirusovgrippaah3n2kotoryecirkulirovalivžitomirskoioblastivperiodépidemičeskogosezona20132014gg AT kyrychukim filogenetičeskiianalizvirusovgrippaah3n2kotoryecirkulirovalivžitomirskoioblastivperiodépidemičeskogosezona20132014gg AT budzanivskaig filogenetičeskiianalizvirusovgrippaah3n2kotoryecirkulirovalivžitomirskoioblastivperiodépidemičeskogosezona20132014gg AT mironenkoap filogenetičeskiianalizvirusovgrippaah3n2kotoryecirkulirovalivžitomirskoioblastivperiodépidemičeskogosezona20132014gg AT radchenkolv filogenetičeskiianalizvirusovgrippaah3n2kotoryecirkulirovalivžitomirskoioblastivperiodépidemičeskogosezona20132014gg AT smutkooyu filogenetičeskiianalizvirusovgrippaah3n2kotoryecirkulirovalivžitomirskoioblastivperiodépidemičeskogosezona20132014gg |
| first_indexed |
2025-11-25T20:36:50Z |
| last_indexed |
2025-11-25T20:36:50Z |
| _version_ |
1850526688932265984 |
| fulltext |
226
ISSN 0233-7657
Biopolymers and Cell. 2015. Vol. 31. N 3. P. 226–232
doi: http://dx.doi.org/10.7124/bc.0008E4
Viruses and Cell
UDC 575.22 + 578.53
Phylogenetic analysis of infl uenza A viruses (H3N2)
circulating in Zhytomyr region during 2013–2014 epidemic season
O. G. Boyalska1, 2, I. M. Kyrychuk3, I. G. Budzanivska1,
A. P. Mironenko4, L. V. Radchenko4, O. Yu. Smutko1
1 ESC "Institute of Biology", Taras Shevchenko National University of Kyiv
64/13, Volodymyrska Str., Kyiv, Ukraine, 01601
2 SI "Zhytomyr regional laboratory center of State Sanitary and Epidemiological Service of Ukraine"
64, V. Berdychivska Str., Zhytomyr, Ukraine, 10002
3 Main Department of State Sanitary and Epidemiological Service of Ukraine in Zhytomyr region
64, V. Berdychivska Str., Zhytomyr, Ukraine, 10002
4 SI "Gromashevsky L.V. Institute of epidemiology and infectious diseases, NAMS of Ukraine"
5, Amosova Str., Kyiv, Ukraine, 03680
vkot71@mail.ru
Aim. To perform phylogenetic analysis of the hemagglutinin (HA) and neuraminidase (NA) genes of
infl uenza A(H3N2) viruses circulating in the Zhytomyr region during 2013–2014 epidemic season. To
make comparison of the HA and NA genes sequences of the Zhytomyr region isolates with the HA and
NA genes sequences of infl uenza viruses circulating in the world. Methods. Laboratory diagnosis was
conducted by real-time polymerase chain reaction (RT-PCR). In this study the sequencing and phyloge-
netic analysis were carried out. Results. For the fi rst time the genes of infl uenza A(H3N2) viruses iso-
lated in the Zhytomyr region during 2013–2014 epidemic season, coding hemagglutinin and neuramini-
dase were compared with their orthologs. According to the results of this comparison the phylogenetic
tree was constructed. Additionally, the amino acid substitutions of the infl uenza viruses circulating in
Ukraine and worldwide were analyzed. Conclusions. The nucleotide sequences of the infl uenza A(H3N2)
viruses genes HA and NA isolated in the Zhytomyr region were identifi ed. Based on the nucleotide se-
quences of HA and NA we constructed the infl uenza virus phylogenetic tree demonstrating that the virus
isolated in the Zhytomyr region was closely related to the Ukrainian isolate from Kharkov and in the
world to the isolates from Germany, Romania, Italy.
K e y w o r d s: A(H3N2) infl uenza viruses, phylogenetic analysis, epidemic season, mutations.
© 2015 O. G. Boyalska et al.; Published by the Institute of Molecular Biology and Genetics, NAS of Ukraine on behalf of Biopolymers and Cell.
This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/),
which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited
Introduction
Infl uenza viruses are the most prevalent pathogens
of human respiratory infections and the most sig-
nifi cant because they cause high morbidity and mor-
tality [1]. Infl uenza affects all age groups of popu-
lation, but the highest incidence is recorded among
children and adolescents [2]. Infl uenza viruses ha-
ve the ability to cause annual epidemy, and someti-
mes – global pandemic. This is applied to the ma-
jority of infl uenza A viruses. These pathogens are
more variable than the infl uenza B and infl uenza C
viruses due to uni que antigenic properties of two
surface glycoproteins: hemagglutinin (HA) and neu-
raminidase (NA) [3].
Infl uenza viruses change the composition of their
surface antigens with high evolutionary rate that al-
lows them to evade the action of the immune system,
and thus to keep themselves in the human population
[4]. There are two main evolutionary mechanisms,
227
Phylogenetic analysis of infl uenza A viruses (H3N2) circulating in Zhytomyr region during 2013–2014 epidemic season
which allow infl uenza viruses to constantly evolve
and re-infect their hosts, namely, an antigenic drift
and antigenic shift [3, 5–7]. The antigenic drift is a
result of the gradual accumulation of mutations that
are fi xed in the viral genome. Such mutations can lead
to minor changes in the viral proteins resulting in bet-
ter survival of these viruses, including the capacity to
escape immune system recognition. During the anti-
genic shift, infl uenza A virus can get segment HA, and
possibly NA segment or another segment of the ge-
nome of different infl uenza virus subtype, resulting in
a radically new infl uenza virus with new properties
[6]. A high variability of infl uenza viruses makes it
necessary to conduct a comparative study of antigenic
and biological properties of infl uenza virus during
epidemics [5]. Also, surveillance and control of anti-
genic properties of circulating infl uenza viruses in the
population are annually required for the defi ning of a
new variant strain for vaccine [2, 8–10].
Thus, the aim of our research was to perform phylo-
genetic analysis of HA and NA genes of infl uenza
A(H3N2) viruses, which circulated in the Zhytomyr
region during the 2013–2014 epidemic season and their
comparison with those which circulated in the world.
Materials and Methods
The material for the study were the clinical samples
of nasopharyngeal swabs, pharyngeal swab, naso-
pha rynx and nose, autopsy material selected in the
fi rst three days and no later than the 5th day of illness
from the patients with suspected infl uenza and SARI
(severe acute respiratory infections) [11]. The pa-
tients were hospitalized in the Zhytomyr region chil-
dren and adult infectious departments of hospitals.
The autopsy material from dead patients was ob-
tained from the regional department of morbid anat-
omy and offi ce forensics.
For carrying out the work there were used viro-
logical and molecular genetic methods of research.
Laboratory diagnosis was conducted by real-time
PCR (polymerase chain reaction). To detect infl uenza
virus types A and B there were used extraction kits
(Total RNA Mini Kit Spin Format, Bio-Rad, USA)
and reagent for reverse transcription (iScript cDNA
Synthesis Kit, Bio-Rad, USA), primers and probes re-
spective markers – Univ inf A, sw A, sw H1, H1, H3,
Univ inf B, RNase P (Biosearch, USA).
The researches were carried out by the protocol (it
was given by WHO) of polymerase chain reaction
with reverse transcription real-time to detect and
study infl uenza A (H1N1) pdm from the Center of
Disease Control and Prevention (CDC) United States
(version 2009) [12].
Virological studies were carried out by the method
of isolation and cultivation of infl uenza viruses in
MDCK cell (epithelial cells kidneys female Cocker-
Spaniel) [10]. The isolates were later used for the
strain identifi cation and sequencing.
The sequencing of infl uenza viruses isolates was
carried out in the WHO Collaborating Centre in Lon-
don. The sequences of infl uenza viruses isolated in ot-
her countries were found using web-site GISAID (the
Global Initiative on Sharing All Infl uenza Data –
http://platfom.gisaid.org). The package of software
MEGA 6.0 for carrying out the phylogenetic analy-
sis was used [13].
Results and Discussion
Infl uenza A (H3N2) virus in the Zhytomyr region
was detected during two epidemic seasons: 2011–
2012 and 2013–2014. In 22 samples (30.5 %) of the
72 samples tested during the epidemic season 2011–
2012 infl uenza A (H3N2) viruses were detected by
PCR in real time [12].
In this study, we have characterized infl uenza
A(H3N2) viruses circulating during 2013–2014 epi-
demic season in the Zhytomyr region.
There were examined 67 nasopharyngeal swabs
from patients with severe course of SARI and sam-
ples of autopsy material. 298 tests by PCR in real
time were carried out. This was done for the moni-
toring of infl uenza viruses circulation among differ-
ent groups of the Zhytomyr region population during
2013–2014 epidemic season. Infl uenza A (H3N2) vi-
ruses were detected in 18 samples (26.9 %). Other
infl uenza viruses in the epidemic season were not
found. The obtained results indicate widespread in-
fl uenza among the different age groups of the
Zhytomyr region, but mostly children and young
adults were involved in the epidemic process.
228
O. G. Boyalska, I. M. Kyrychuk, I. G. Budzanivska et al.
Fig. 1. Phylogenetic analysis of the HA gene of infl uenza A (H3N2) viruses isolated in the Zhytomyr region during 2013–2014 epi-
demic season
A/Ukraine/6101/2014
A/Ukraine/77/2014
A/Belgrade/2668/2014
A/England/256/2014
A/Finland/387/2013
A/Serbia/NS-669/2014
A/Bratislava/111/2014
A/Macedonia/IK (80)/2014
A/Kharkov/203/2014
A/Zaporizza/324/2014
A/Denmark/103/2013
A/Bucuresti/161525/2014
A/Baden-Wurttemberg/1/2014
A/Zhitomir/290/2014
A/Zhitomir/286/2014
A/Kharkov/201/2014
A/Parma/3/2014
A/Bulgaria/800/2014
A/Georgia/328/2014
A/Ankara/221/2014
A/Hawaii/09/2014
A/Korea/3042/2014
A/Alaska/08/2014
A/Serbia/NS-210/2013
A/South Africa/4655/2013
A/Samara/73/2013
A/Almaty/2958/2013
A/Dnipro/229/2014
A/Ukraine/6138/2014
A/Ukraine/728/2013
A/Croatia/2561/2013
A/Hong Kong/146/2013
A/Ukraine/6161/2014
A/Georgia/495/2014
A/Dnipro/234/2014
A/Latvia/03-013033/2014 A/SYDNEY/20/2014
A/New York/07/2014
A/Stockholm/1/2013
A/Moscow/206/2014
A/Israel/Z125/2013
A/Vietnam/480/2013
A/Slovenia/525/2014
A/Poland/3464/2014
A/Khmelnitsky/249/2014
A/Belgium/14G0001/2014
A/Victoria/361/2011
A/Texas/50/2012
98
67
98
99
80
68
60
91
96
76
75
67
63
98
88
89
71
64
69
57
81
0.002
The vaccine strain
Reference strains
Ukrainian isolates
Zhytomyr region
isolates
K27R
N122D
V529I
D489N
L157S
V347M I214T
T128A
R142G
N225D
K326R
N145S
D489N
Y94H
N225D
Q311H
V402I
G78D
A304D
T128V
1
2
I242V P289S
S312N Q33R
N145S
N278K
229
Phylogenetic analysis of infl uenza A viruses (H3N2) circulating in Zhytomyr region during 2013–2014 epidemic season
For isolation of infl uenza viruses in MDCK cell
culture we used fi ve samples, in which infl uenza
A(H3N2) viruses were detected [10].
For comparison, in our research we used the seg-
ments that encode the surface proteins HA and NA.
These proteins are mostly exposed to antigenic drift
and shift to avoid the effects of human immune sys-
tem. Hemagglutinin is a receptor-binding protein that
is the main target of the immune response. Therefore
it is the most variable protein in infl uenza viruses as
amino acid substitutions in the antigenic sites (A, B,
C, D, E) of HA molecules affect the antigenic prop-
erties of infl uenza virus. These events infl uence the
change of vaccine strains. He The neuraminidase
protein is less variable, but amino acid substitutions
in it affect the resistance of infl uenza viruses to the
antiviral drugs (neuraminidase inhibitors) [14].
The isolates were sent to the World Infl uenza Cen-
tre (London) for sequencing. In the epidemic season
2013–2014 for the fi rst time there were made the se-
quences of genes HA and NA of infl uenza A(H3N2)
viruses isolated from the clinical material from pa-
tients of the Zhytomyr region and they were subse-
quently included in the GISAID database of infl u-
enza viruses worldwide.
We found a very high similarity (99 %) of the HA
and NA genes of the Zhytomyr region viruses with the
HA and NA genes of infl uenza viruses in the group
with T214I substitution on the phylogenetic tree using
BLAST system. Comparison of the nucleotide se-
quences of the Zhytomyr region virus HA and NA with
those of the T214I group viruses revealed several syn-
onymous nucleotide substitutions and no substitutions
that alter amino acid sequences of the proteins.
The HA and NA phylogenetic tree of the Zhytomyr
region strains and strains from all over the world was
generated using MEGA 6.0. Evolutionary distance
was calculated using the maximum composite likeli-
hood method. Statistical signifi cance of the tree to-
pology was tested by bootstrap analysis of 100 pseu-
doreplicate datasets.
The 2728 HA and NA protein sequences of infl u-
enza A(H3N2) viruses of different strains circulated
during 2013–2014 epidemic season in the world we-
re downloaded from the GISAID database.
By random sampling we selected 48 isolates from
around the world during 2013–2014 epidemic sea-
son and constructed the phylogenetic tree of HA
genes (Fig. 1).
All the HA genes of the Zhytomyr region isolates,
as all Ukrainian and worldwide isolates studied with
vaccine strain A/Texas/50/2012 and reference strains:
A/Victoria/361/2011, A/Hong Kong/146/2013, A/
Stockholm/1/2013, A/Serbia/NS-210/2013, A/South
Africa/4655/2013, A/Samara/73/2013, A/Almaty/2958/
2013 fell within the A/Victoria/208 genetic and into
the of genetic subgroup 3C of group 3 [15]. All sub-
mitted isolates acquired in the evolution new joint
(Q33R), group (L157S, I214T, N122D) and individ-
ual (V402I) amino acid substitution in a part of he-
magglutinin compared with vaccine strain and refer-
ence strains. Therefore, they can be divided into two
groups. The amino acid substitutions that defi ne the-
se groups are:
The fi rst genetic group included the isolates that
had substitution N145S (asparagine to serine) and
D489N (aspartic acid to asparagine). The group com-
bined some viruses from Ukraine and the world and
reference strains A/Hong Kong/146/2013 and A/Sto-
ckholm/1/2013.
In turn, the reference strain A/Stockholm/1/2013 has
substitution I242V and P289S, and virus A/Hong Kong/
146/2013 acquired a new glycosylation site S312N.
The second genetic group comprised a large quan-
tity of Ukrainian viruses including the Zhytomyr re-
gion isolates and reference strains that had T128A
(resulting in the loss of a potential glycosylation si-
te), R142G substitutions, along with the isolates from
all over the world. The group combined some viruses
from Ukraine, the world and reference strains A/Ser-
bia/NS-210/2013, A/So uth Africa/4655/2013, A/Sa-
ma ra/73/2013, A/Almaty/2958/2013.
The Zhytomyr region isolates were found to be
most closely related to the Ukrainian isolate from
Khar kov as well as to the isolates from Germany,
Ro mania, Italy. They have gained substitution I214T
(isoleucine to threonine).
Thus, analyzing the HA gene sequences of the
Zhytomyr isolates we can conclude that the strains
recorded in the Zhytomyr region, are similar to those
230
O. G. Boyalska, I. M. Kyrychuk, I. G. Budzanivska et al.
A/Victoria/361/2011
A/Texas/50/2012
A/Korea/3302/2014
A/Hawaii/09/2014
A/Alaska/06/2014
A/SYDNEY/120/2014
A/Georgia/02/2014
A/Washington/13/2014
A/Romania/157360/2014
A/Kaliningrad/01/2014
A/Ankara/221/2014
A/Latvia/03-013033/2014
A/Georgia/495/2014
A/Kharkov/203/2014
A/Slovenia/108/2014
A/Belgium/14G0001/2014
A/Stockholm/1/2013
A/Moscow/206/2014
A/Milano/113/2014
A/Hong Kong/146/2013
A/Samara/73/2013
A/Norway/58/2014
A/Serbia/NS-210/2013
A/Latvia/12019591/2013
A/Saint-Petersburg/RII03/2014
A/Netherlands/2247/2013
A/South Africa/5416/2014
A/South Africa/4655/2013
A/Croatia/2561/2013
A/Hong Kong/33/2014
A/Dnipro/384/2014
A/Slovenia/1213/2014
A/Dnipro/227/2014
A/Ukraine/728/2013
A/Ukraine/6097/2014
A/Almaty/2958/2013
A/Athens GR/112/2012
A/Zhitomir/290/2014
A/Zhitomir/286/2014
A/Kharkov/201/2014
A/Baden-Wurttemberg/1/2014
A/Serbia/NS-682/2014
A/Bulgaria/827/2014
A/Stockholm/15/2014
A/Madagascar/00153/2014
A/Ukraine/222/2014
A/Ukraine/218/2014
A/Belgrade/2668/2014
A/Dnipro/232/2014
A/Ukraine/77/2014
A/Milano/84/2014
A/Ukraine/273/2014
A/Khmelnitsky/244/2014
A/Poland/896/2014
A/Hawaii/02/2014
A/Norway/1003/2014
A/Israel/Z774/2014
A/Hong Kong/5383/2014
A/Macedonia/IK (80)/2014
A/Zaporizza/324/2014
A/Denmark/103/2013
A/Georgia/76/2014
A/Bratislava/111/2014
A/Togo/LNG/398/2013
A/Perth/16/2009
80
84
56
58
91
90
92
91
82
66
71
80
61
55
43
93
99
87
67
69
47
47
66
71
60
63
42
100
90
0,0001
The vaccine strain
Reference strains
Ukrainian isolates
Zhytomyr region
isolates
1
T434I
T267KE221D
E221D V412I I65V N141D
2
S367N
K369T
L81P
N402D
Y155F
D251V
S315G
3
A246D
T434I
R210K
Q5K S332F
N329T I307T
I392T
I312T
Fig. 2. Phylogenetic analysis of the NA gene of infl uenza A (H3N2) viruses isolated in the Zhytomyr region during 2013–2014 epi-
demic season
231
Phylogenetic analysis of infl uenza A viruses (H3N2) circulating in Zhytomyr region during 2013–2014 epidemic season
from other regions of Ukraine (the Kharkiv region)
or from abroad (Germany, Romania, Italy). This is
due to a high level of the migration both within the
country and abroad.
In the phylogenetic tree of the NA genes 65 iso-
lates from around the world were presented (Fig. 2).
The analysis of the NA genes of the Zhytomyr re-
gion infl uenza viruses showed similar results with
the analysis of the HA gene sequences. All investi-
gated isolates are within the genetic subgroup 3C of
A/Victoria/208 genetic clade, keeping mutation S367N
and N369T. These mutations are typical for the group
3. The mutations L81P and N402D (loss glycosyla-
tion site) are typical for the subgroup 3C.
All studied isolates of the 2013–2014 epidemic
season were genetically similar to the vaccine strain
A/Texas/50/2012, but had some genetic differences,
so they were divided into three genetic groups.
The fi rst group includes the viruses from differ-
ent countries including Ukraine – A/Kharkov/203/
2014 (V396I) with substitution I392T (isoleucine
to threonine).
The second group is based on the Ukrainian iso-
lates and the isolates from different countries of the
world with replacement E221D (glutamic acid to as-
partic acid substitution) and V412I (valine to isoleu-
cine substitution).
Our isolates were in the group which included the
Ukrainian isolates and the isolates from around the
world. In this group there were found new substitu-
tions Y155F, D251V, S315G. Our isolates are locat-
ed along with the viruses from Kharkiv and Baden-
Württemberg. This group is very heterogeneous and
is similar to the reference virus A/Athens GR / 112/
2012 more than to new reference viruses.
Conclusions
The Zhytomyr region isolates were found to be most
closely related to the Ukrainian isolate from Kharkov
and in the world to the isolates from Germany,
Romania, Italy. All these viruses are specifi ed by the
substitution I214T. Our infl uenza viruses of 2013–
2014 epidemic season have a high genetic relation-
ship (99 %) to the viruses carrying the I214T substi-
tutions. All nucleotide substitutions in the genomes
of the Zhytomyr region isolates are synonymous.
The Zhytomir region isolates were placed in a domi-
nant subgroup world 3C of A/Victoria/208 genetic
clade. They, like many Ukrainian and world isolates,
were similar to the reference strain A/AthensGR/
112/2012, more than to new reference viruses.
Acknowledgements
The authors thank Oleksandr Volkov (Chief of Main
Department of State Sanitary and Epidemiological
Service of Ukraine in Zhytomyr region), Zynoviy
Paramonov (SI «Zhytomyr regional laboratory cent-
er of State Sanitary and Epidemiological Service of
Ukraine») and World Infl uenza Centre (London,
Great Britain) for help in carrying out sequencing
the Zhytomyr region isolates.
REFERENCES
1. Boyalska O, Kyrychuk I, Shpyta O, Boyko A. Dynamics of
fl ue morbidity among the population of Zhytomyr region.
Agroecologichnyy Zhurnal. 2014; 4: 106–9.
2. Campa A, Quattrocchi M, Guido M, Gabutti G, Germinario
C, De Donno A, Group TI. Ten-year (1999–2009) epidemio-
logical and virological surveillance of infl uenza in South
Italy (Apulia). Infl uenza Res Treat. 2010;2010:642492.
3. Zhang S, Chen JY, Rao J, Zhang S, Wai Chan DWT, Liu G,
Xu Y, He M. Genetic variation analysis of hemagglutinin
and neuraminidase of human infl uenza A(H3N2) virus in
Hong Kong (1997–2006). Afr J Microbiol Res. 2014; 8(18):
1863–93.
4. Rickert JD, Herpy TA, Zhong H, Lu G. Computational anal-
ysis of adaptive antigenic mutations of the human infl uenza
hemagglutinin for vaccine strain selection. Int J Bioinform
Res Appl. 2012;8(1–2):81–98.
5. Stray SJ, Pittman LB. Subtype- and antigenic site-specifi c
differences in biophysical infl uences on evolution of infl u-
enza virus hemagglutinin. Virol J. 2012;9:91.
6. Escalera-Zamudio M, Nelson MI, Cobián Güemes AG, Ló-
pez-Martínez I, Cruz-Ortiz N, Iguala-Vidales M, García ER,
Barrera-Badillo G, Díaz-Quiñonez JA, López S, Arias CF,
Isa P; Members of Colegio de Pediatría del Estado de Ve ra-
cruz. Molecular epidemiology of infl uenza A/H3N2 viruses
circulating in Mexico from 2003 to 2012. PLoS One. 2014;
9(7):e102453.
7. Zhong J, Liang L, Huang P, Zhu X, Zou L, Yu S, Zhang X,
Zhang Y, Ni H, Yan J. Genetic mutations in infl uenza H3N2
viruses from a 2012 epidemic in Southern China. Virol J.
2013;10:345.
8. Bedford T, Suchard MA, Lemey P, Dudas G, Gregory V, Hay
AJ, McCauley JW, Russell CA, Smith DJ, Rambaut A. In te-
232
O. G. Boyalska, I. M. Kyrychuk, I. G. Budzanivska et al.
grating infl uenza antigenic dynamics with molecular evolu-
tion. Elife. 2014;3:e01914.
9. DE Donno A, Idolo A, Quattrocchi M, Zizza A, Gabutti G,
Romano A, Grima P, Donatelli I, Guido M. Surveillance of
human infl uenza A(H3N2) virus from 1999 to 2009 in so ut-
hern Italy. Epidemiol Infect. 2014;142(5):933–9.
10. WHO Global Infl uenza Surveillance Network Manual for
the laboratory diagnosis and virologic al surveillance of in-
fl uenza. 2011; 153p.
11. Boyalska OG, Kyrychuk IM, Mironenko AP, Radchenko LV.
Virological infl uenza and other ARVI surveillance in the
Zhytomyr Region. Visnyk Problem Biologii ta Meditsyny.
2015; 3(120)(2): 95–100.
12. World Health Organization (WHO). CDC protocol of real-
time RT-PCR for infl uenza H1N1. World Health Orga ni za-
tion, Geneva, Switzerland. 30 April 2009.
13. Tamura K, Stecher G, Peterson D, Filipski A, Kumar S.
MEGA6: Molecular Evolutionary Genetics Analysis ver-
sion 6.0. Mol Biol Evol. 2013;30(12):2725–9.
14. Eshaghi A, Duvvuri VR, Li A, Patel SN, Bastien N, Li Y, Low
DE, Gubbay JB. Genetic characterization of seasonal infl u-
enza A (H3N2) viruses in Ontario during 2010–2011 infl u-
enza season: high prevalence of mutations at antigenic sites.
Infl uenza Other Respir Viruses. 2014;8(2):250–7.
15. Report prepared for the WHO annual consultation on the
composition of infl uenza vaccine for the Southern He mis p-
here 2015 (22–24 September 2014). WHO Collaborating
Centre for Reference and Research on Infl uenza National
Institute for Medical Research The Ridgeway Mill Hill
London, NW7 1AA. 2014; 38–85.
Філогенетичний аналіз вірусів грипу
А(H3N2), що циркулювали в Житомирській області
протягом епідемічного сезону 2013–2014рр.
О. Г. Бояльська, І. М. Киричук, І. Г. Будзанівська,
А. П. Міроненко, Л. В. Радченко, О. Ю. Смутько
Мета. Зробити філогенетичний аналіз генів гемаглютиніну
(HA) і нейрамінідази (NA) вірусів грипу А(H3N2), що були
поширені в Житомирській області під час епідемічного се-
зону 2013–2014рр. і порівняти їх з тими, що циркулювали в
світі. Методи. Лабораторну діагностику здійснювали за до-
помогою ПЛР у реальному часі (real-time RT-PCR). Було
проведено секвенування та філогенетичний аналіз.
Результати. В епідемічному сезоні 2013–2014 рр. вперше
було зроблено секвенування генів НА та NA вірусів грипу
А(H3N2), виділених з клінічного матеріалу хворих з
Житомирської області та в подальшому увійшли до бази да-
них вірусів грипу з усього світу (GISAID). Висновки.
Житомирські ізоляти розташувались поряд з українським
ізолятом з Харкова, а серед світових поряд з ізолятами з
Німеччини, Румунії, Італії. В послідовностях генів гемаглю-
тиніну було виявлено заміщення I214T. Виділені нами ізо-
ляти в епідемічному сезоні 2013–2014рр. мали високу гене-
тичну спорідненість (99 %) до вірусів, що знаходяться в
групі на філогенетичному дереві, тому що мають синоніміч-
не заміщення. Житомирські ізоляти розташувались у домі-
нуючій групі в світі 3С групи 3 генетичного кластеру A/Vic-
toria/208 і були подібними до вакцинного штаму A/Texas/
50/2012. В послідовностях генів нейрамінідази суттєвих за-
міщень виявлено не було. Житомирські ізоляти, як і багато
українських та світових ізолятів виявились подібними до
референс-штаму A/AthensGR/112/2012, ніж до більш нових
референс-вірусів.
Ключов і слова: віруси грипу A(H3N2), філогенетичний
аналіз, епідемічний сезон, мутації.
Филогенетический анализ вирусов гриппа А(H3N2)
которые циркулировали в Житомирской области
в период эпидемического сезона 2013–2014гг.
О. Г. Бояльская, И. Н. Киричук, И. Г. Будзанивская,
А. П. Мироненко, Л. В. Радченко, О. Ю. Смутько
Цель. Провести филогенетический анализ генов гемагглю-
тинина (HA) и нейраминидазы (NA) вирусов гриппа А(H3N2),
которые циркулировали в Житомирской области во время
эпидемического сезона 2013–2014гг. и сравнить их с тако-
выми со всего мира. Методы. Лабораторную диагностику
проводили с помощью ПЦР в реальном времени (real-time
RT-PCR). Было проведено секвенирование и филогенети-
ческий анализ. Результаты. В эпидемическом сезоне 2013–
2014гг. впервые было сделано секвинирование генов НА та
NA вирусов гриппа А(H3N2), виделенных из клинического
материала больных в Житомирськой области и в дальней-
шем вошли в базу данных вирусов гриппа со всего мира
(GISAID). Выводы. Житомирские изоляты разместились
рядом с украинским изолятом из Харькова, а среди мировых
рядом с изолятами из Германии, Румынии, Италии. В после-
довательностях генов гемагглютинина было выявлено заме-
щение I214T. Выделенные нами изоляты в эпидемическом
сезоне 2013–2014гг. имели высокое генетическое сходство
(99 %) к вирусам, которые находяться в группе на филогене-
тическом дереве, так как имеют синонимическое замеще-
ние. Житомирские изоляты разместились в доминирующей
субгруппе в мире 3С группы 3 генетического кластера A/
Victoria/208. В последовательностях генов нейраминидазы
существенных мутаций не выявлено. Житомирские изоля-
ты, как много украинских и мирових изолятов оказались
подобными к референс-штамму A/AthensGR/112/2012, чем
к более новым референс-вирусам.
Ключевые слова: вирусы гриппа A(H3N2), филогене-
тический анализ, эпидемический сезон, мутации
Received 07.02.2015
|