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<title>Genetic variability of attachment (G) and Fusion (F) protein genes of human metapneumovirus strains circulating during 2006-2009 in Kolkata, Eastern India</title>
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Agrawal et al. Virology Journal 2011, 8:67
http://www.virologyj.com/content/8/1/67
RESEARCH
Open Access
Genetic variability of attachment (G) and Fusion
(F) protein genes of human metapneumovirus
strains circulating during 2006-2009 in Kolkata,
Eastern India
Anurodh S Agrawal, Tapasi Roy, Swati Ghosh, Mamta Chawla-Sarkar*
Abstract
Background: Human metapneumovirus (hMPV) is associated with the acute respiratory tract infection (ARTI) in all
the age groups. However, there is limited information on prevalence and genetic diversity of human
metapneumovirus (hMPV) strains circulating in India.
Objective: To study prevalence and genomic diversity of hMPV strains among ARTI patients reporting in
outpatient departments of hospitals in Kolkata, Eastern India.
Methods: Nasal and/or throat swabs from 2309 patients during January 2006 to December 2009, were screened
for the presence of hMPV by RT-PCR of nucleocapsid (N) gene. The G and F genes of representative hMPV positive
samples were sequenced.
Results: 118 of 2309 (5.11%) clinical samples were positive for hMPV. The majority (≈80%) of the positive cases
were detected during July−November all through the study period. Genetic analysis revealed that 77% strains
belong to A2 subgroup whereas rest clustered in B1 subgroup. G sequences showed higher diversity at the
nucleotide and amino acid level. In contrast, less than 10% variation was observed in F gene of representative
strains of all four years. Sequence analysis also revealed changes in the position of stop codon in G protein, which
resulted in variable length (217-231 aa) polypeptides.
Conclusion: The study suggests that approximately 5% of ARTI in the region were caused by hMPV. This is the first
report on the genetic variability of G and F gene of hMPV strains from India which clearly shows that the G
protein of hMPV is continuously evolving. Though the study partially fulfills lacunae of information, further studies
from other regions are necessary for better understanding of prevalence, epidemiology and virus evolution in
Indian subcontinent.
Background
Acute Respiratory tract infections (ARTI) are a leading
cause of morbidity and mortality worldwide [1]. Human
metapneumovirus (hMPV), genus Metapneumovirus,
family paramyxoviridae first identified in the Netherlands [2], is an important etiological agent of acute
respiratory tract infection in almost all age groups. Subsequently it has been identified all over the world [3-6].
Morphologically, hMPV consists of a negative-sense,
* Correspondence: chawlam70@gmail.com
Division of Virology, National Institute of Cholera and Enteric Diseases, P-33,
C.I.T. Road Scheme XM, Beliaghata, Kolkata-700010, India
single stranded and non Segmented RNA that encodes
at least 9 distinct proteins [7]. Among them, the two
major transmembrame glycoproteins, G and F, stimulate
the production of protective immune responses, and
therefore, are antigenically significant [8]. F protein promotes fusion of the viral and cell membrane while G
protein mediates virus binding to the cell receptor [9].
Genetic analysis on the basis of N, M and F genes
have classified hMPV into two distinct groups or genotypes A and B [10-13]. Both genotypes are known to be
prevalent throughout the world and circulate in a single
season with the switching of predominant group in
© 2011 Agrawal et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons
Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in
any medium, provided the original work is properly cited.
Agrawal et al. Virology Journal 2011, 8:67
http://www.virologyj.com/content/8/1/67
successive seasons [3,12,14-16]. Unlike the relatively
conserved F protein (95% identity at the amino acid
level between group A and B), the G protein is highly
variable with only 53% amino acid homology between
group A and B [17,18].
In developing countries like India, approximately 0.5
million children <5 years of age die due to ARTI
[19-21]. We have previously reported prevalence of
Influenza A (11±1%), influenza B (5.5± 0.5%) and RSV
(7.5 ±1%) among outdoor patients in Kolkata [22,23].
Inspite of its significance as an important respiratory
pathogen, there is no information on prevalence and
genetic diversity of hMPV strains in India except for
one report from Northern India [24]. To partially fulfill
this lacuna, the study was done to analyze the extent of
genetic variation and the circulation pattern of hMPV in
Kolkata during 2006-2009.
Methods
Sampling site and Study Population
The study was conducted among patients of all age group
exhibiting fever and 2 or more symptoms of ARTI (cold/
cough, sore throat, myalagia, body ache) from the outdoor patient ward of hospitals in Kolkata as reported previously [23]. None of these patients were hospitalized.
Nasal and/or throat swabs were collected from 2309
patients and were transported in viral transport media
(VTM) to the laboratory. The study was approved by the
Institutional Ethical Committee and the informed consent was taken from patients or their guardians.
Page 2 of 8
[26,27]. The multiple and pair wise alignment of
deduced amino acid (aa) sequences were performed by
using CLUSTAL W software and phylogenetic trees
were generated by the neighbor-joining method with the
MEGA 5 software as described earlier [28].
Nucleotide sequence database accession numbers
The hMPV sequences for the 22 G and 8 F genes analyzed in this study have been deposited in GenBank
under the accession number HQ599198-HQ599227.
Results
Prevalence & age distribution of hMPV
A total of 2309 samples were screened during January
2006 to December 2009 by RT-PCR based amplification
of the relatively conserved N gene. Although the age of
the patients ranged from 1 month to 50 years, most
(≥78%) were below 5 years of age (Table 1). The screening results were confirmed by sequencing and BLAST
analysis of N gene amplicon. hMPV was identified in
118 (5.11%) samples in four years. 32 (6.32%) of the 506
samples in 2006, 27 (4.46%) of 605 samples in 2007, 41
(5.84%) of 702 samples in 2008 and 18 (3.63%) of 496
samples in 2009 were positive for hMPV.
The hMPV positive samples were found at low frequency (0.5-1%) throughout the year but the majority
(≈80%) of the hMPV positive samples were detected
during July−November correlating positively with rainfall and high humidity.
Phylogenetic and antigenic analysis of G protein
Extraction of viral RNA
RNA was extracted from 200 ul clinical samples using
commercially available RNeasy Mini Kit (Qiagen GmbH,
Hilden, Germany) as per manufacturer’s instructions.
Reverse transcription and PCR
For initial screening, amplification of a 416 bp portion
of nucleoprotein (N) gene was carried out using primers
hmpv1 and hmpv2 by RT-PCR as described earlier [25].
All N positive samples were further amplified by using
previously described G and F gene specific primers
[12,24]. The resulting PCR products were purified with
a Qiagen PCR purification Kit.
Sequence and sequence analysis
Nucleotide (nt.) sequencing of full length G gene and
partial F gene (nt.1-nt.805) was carried out by using ABI
Prism Big Dye Terminator v3.1 Cycle Sequencing Ready
Reaction Kits in an ABI Prism 3100 Genetic Analyzer
(PE Applied Biosystems, Foster City, California, U.S.A)
using gene specific forward and reverse primers. Potential N’- and/or O’- glycosylation site/s were predicted by
using NetNGlyc 1.0 and NetOGlyc v.3.1 software
Phylogenetic analysis of 22 Kolkata strains (five representative strains from each year), confirmed two main
genetic lineages A and B. Each lineage A and B was
further divided into 2 sub-lineage A1 & A2 and B1 &
B2 respectively (Figure 1). Interestingly, all the Kolkata
strains clustered with A2 and B1 sub-lineage only (Figure 1). During 2006 and 2007, both sub group A2 and
B1 co-circulated, with 77% (n = 59) of the circulating
strains belonging to A2 subgroup. Of 32 hMPV positive
samples in 2006, 26 were as subgroup A2 and 6 as subgroup B1, whereas in 2007, 27 were A2-positive and 8
were B1 positive strains. Interestingly no B1 strains were
found in 2008 and 2009 and subgroup A2 remained as
dominant strain throughout the study.
Sequence analysis of the complete ORF of G protein,
revealed homology ranging from 53.8- 56.4% at nt level
Table 1 Prevalence of hMPV infection in different age
groups among outpatients (n = 2309)
Age Group/
Virus prevalence
0-1 year
n = 449
1-2 year
n = 782
2-5 yr
n = 632
≥ 15 yr
n = 446
hMPV
4.23%
5.37%
7.28%
2.47%
Agrawal et al. Virology Journal 2011, 8:67
http://www.virologyj.com/content/8/1/67
Page 3 of 8
Figure 1 Phylogenetic analysis of the complete G gene of 22 hMPV Kolkata strains. Phylogenetic analysis of nt. sequences of hMPV group
A and B strains from Kolkata (named with the prefix KOL followed by sample number and the year of collection) with that of other hMPV strains
from different subgroups. Trees were built using neighbor-joining algorithm through MEGA 4 program. The tree was rooted with cognate
stretch of G gene of strain AMPV-C (GenBank accession number AY198394).
and 34.1-35.9% at the aa level between the members of
group A and B isolates. Sub group B1 strains shared
high percentage of homology with the prototype strain
NL/1/99 {91.8%−92.6% (nt.); 89.1%−90.0% (aa)}, whereas
subgroup A2 strains revealed homology with the prototype strain CAN97-83 {84.5%−90.8% (nt.); 76%-86.3%
(aa)}.
The alignment of deduced aa sequence of G protein of
Kolkata strains with their prototype strains revealed that
intracellular and transmembrane regions were highly
conserved across the strains (Figure 2 and Figure 3).
Most of the aa changes were observed in extracellular
domain due to nt substitution and insertion. Changes in
position of stop codon have been observed among
strains of different subgroups {nt 658 (UAG); nt 652
(UAA), nt 685 (UAG), nt 694 (UGA)}, which correspond
to variable lengths in polypeptides. Strains from subgroup A2 used two different stop codons resulting in G
proteins of 217aa (UAA), 219aa (UAG) and 228aa
(UAG) (Figure 2), whereas subgroup B1 strains
Agrawal et al. Virology Journal 2011, 8:67
http://www.virologyj.com/content/8/1/67
Page 4 of 8
Figure 2 Deduced amino acid sequence of G ORF of hmpv Group A strains. Multiple alignment of aa sequences of G protein gene of 17
hMPV subgroup A2 strains from Kolkata. The prototype strain CAN97-83 (GenBank accession number AY485253) were considered as
representative for group A. Identical residues are indicated by dots and dashes represent gaps. Cysteine residues are marked with asterisks.
Potential N-glycosylation sites are underlined.
Agrawal et al. Virology Journal 2011, 8:67
http://www.virologyj.com/content/8/1/67
Page 5 of 8
Figure 3 Multiple alignment of aa sequences of G protein gene of hMPV group B strains. Deduced aa sequence of complete G ORF of 5
hMPV subgroup B1 strains from Kolkata. The prototype strain NL/1/99 (GenBank accession number AY525843) were considered as representative
for group B. Identical residues are indicated by dots and dashes represent gaps. Cysteine residues are marked with asterisks. Potential Nglycosylation sites are underlined.
terminated at UGA stop codon and exhibited protein of
231 aa in length (Figure 3). For both the subgroup A2
and B1, a cysteine residue at position 27 is strictly conserved among all isolates in the intracellular domain
except in one strain Kol/1446/08 which did not contain
any cysteine residue. The G ectodomain also has a high
content of proline residues, ranging from 7.8% for group
B to 10% for group A, which could contribute to an
extended, unfolded secondary structure.
The G protein gene sequenced in this study exhibited
high content of serine and threonine residues that are
potential O-linked sugar acceptors in both subgroups
A2 and B1. Serine and threonine content of group A
and group B strains was in the range 34.2−37.72% and
29.4−31.18% respectively. The program NetOglyc v. 3.1
predicted 45 to 55 serine and threonine residues to be
potentially O-glycosylated with score predictors (G
scores) of between 0.5 and 0.8. All the predicted O-glycosylation sites were located in the extracellular region
of the subgroup A2 and B1.
The number of N-linked glycosylation site present in
the G protein from different subgroup varied from two
to six, and only one conserved site (aa 30) at the junction of the intracellular and transmembrane domain
[10,29]. The rest of the sites showed subgroup specific
conservation: sites 101, 169, 181 & 188 were conserved
in all the B1 strains whereas site 52, 145 and 152 was
conserved among all the strains of subgroup A2. The
predicted N-linked glycosylation sites at aa 52 (subgroup
A2) and aa 30 (subgroup B1) exhibited high score of 0.7.
Only strain Kol/30/06 lacked the potential site at aa 52
whereas Kol/1367/08 and Kol/2075/09 had lost sites at
aa 145 and 152.
Analysis of the F-gene
Out of 118 hMPV positive Kolkata strains, F-gene was
partially sequenced from positive patients covering
throughout the study period. Blast analysis and sequence
alignment revealed very little difference (≥98% homology) among the strains. Thus the phylogenetic analysis
of the F-gene fragment was done with only 8 representative Kolkata strains (two strains per year). The Kolkata
strains clustered with A2 (six strains) and B1 (two
strains) sub-lineage strains NL/17/00 and NL/1/99
respectively (Figure 4). At the nt level, Kolkata strains
shared higher percentage of homology with subgroup
B1 prototype strain NL/1/99 (98.1%) than the A2 subgroup strain CAN97-83 (96.05−96.8%). Amino acid
alignment of eight partial hMPV F gene (295 aa long)
was compared with the prototype strains from Canada
and the Netherland (see Additional file 1). For both the
subgroup cysteine residues were conserved at position
28, 60, 182, 283 and 292 which could be involved in
proper folding of F monomer, as been suggested for
Agrawal et al. Virology Journal 2011, 8:67
http://www.virologyj.com/content/8/1/67
Page 6 of 8
Figure 4 Phylogenetic analysis of the partial F gene of 8 hMPV Kolkata strains. Phylogenetic analysis of nt. sequences of hMPV group A
and B strains from with that of other hMPV strains. Trees were built using neighbor-joining algorithm through MEGA 4 program. Strains from
different subgroups were included in analysis.
hRSV [30]. Some of the important aa changes were subgroup specific which differentiated group A from group
B (Table 2).
Discussion
In developing countries like India, the mortality and
morbidity risk due to ARTI can be 30 times higher than
in developed countries [31]. In spite of its importance,
very few reports on etiology of ARTI cases are available
[22-24]. hMPV is an important cause of ARI, which has
been found in both healthy and immunocompromised
patients [32,33]. The present study (2006-2009) provided
vital insights into the epidemiology and genomic diversity of hMPV strains circulating among patients in Kolkata city, eastern India. To our knowledge, this is the first
report on genetic diversity of hMPV strains based on G
and F gene sequences from India.
To detect hMPV, initially N gene was chosen as it is
highly conserved and has been used in previous studies
[33,34]. RT-PCR based detection revealed a significant
rate (118/2309) of infection among outpatients with
ARI. Of 118 positives, two samples had dual infection
with RSV though no differences in clinical symptoms
were observed. Compared with Influenza A (11±1%) and
Table 2 List of subgroup specific amino acid changes in the representative Kolkata strains, isolated from 2006-2009
Position/
Subgroup
61
122
135
139
A2
A
B1
T
143
167
175
V
T
I
N
179
185
233
286
N
K
D
G
Q
E
296
R
K
D
N
V
K
S
R
A
Y
I
N
Agrawal et al. Virology Journal 2011, 8:67
http://www.virologyj.com/content/8/1/67
RSV (7.5 ±1%), an average 5.11% (4.6%- 6.3%) positivity
of hMPV was observed among the same study group
[22,23]. This is significantly higher compared to reports
from Canada (2.3%), England (2.2%) and USA (4.5%)
[4,35,36], but is lower compared to frequency reported
from the Netherlands (10%), Australia (9.7%) and Chile
(5.4%) [2,16,37]. These variations in detection rates
could be attributed to factors such as study population,
seasonality and methods for detection. Majority of
hMPV was detected from July to Nov (monsoon and
autumn), which is similar to previous reports [32,38],
but contrary to reports from New Delhi, India and temperate countries [6,24,39], where the high incidence was
observed in cold season. This is consistent with seasonality of influenza viruses which follow different seasonality between tropical and temperate countries [40].
Due to similarity between RSV and hMPV, we analyzed
genetic diversity of both F and G surface glycoproteins
because i) they are two major targets for neutralizing and
protective immunity in RSV [8], ii) hMPV F gene is the
major antigenic determinant and is classified worldwide
into the context of genetic lineages [5,35,41], iii) the G
protein has been described as the most variable gene product among hMPV like the G protein of RSV [11,18,29].
Phylogenetic analysis based on nt sequences of G and F
gene with the representative strains demonstrated the
existence of two group (A and B) and two subgroup A2
and B1. The study demonstrated high prevalence of subgroup A2 (77%) than subgroup B1 (23%) infection. Both
group A and B viruses co-circulated for year 2006-07 but
later group B virus disappeared. Similar to previous studies [11,29], the G gene sequence alignment showed
extensive nt (53.8−56.1%) and aa (34.2−35.9%) variation
between these two groups. In addition different length of
G polypeptide in strains belonging to different subgroup
was also observed due to the usage of different stop
codon [5,42,43]. Further studies are required to know
whether the changes in stop codon are lineage specific
and/or associated with the emergence of new evolutionary lineages, as suggested for RSV [44-46]. On the other
hand, accumulation of sporadic aa substitutions and presence of additional and/or absence of N-’and/or O’- glycosylation sites in subgroup A2 and B1 strain from
Kolkata provided evidences for constant mutation events
which could be either critical for evading immune
response/s or may confer enhanced stability that favor
gradual establishment of certain local strains over others.
Conclusion
Since there was only one report on genetic heterogeneity
of hMPV strains from northern India, it was extremely
difficult to assess the current status of heterogeneity of
hMPV strains in the country. For assessing prevalence,
susceptible age group, and genetic variation, analysis of
Page 7 of 8
hMPV was initiated in addition to other respiratory
pathogens in Eastern India. Even though the information
is not representative for Indian subcontinent, this study
provides the much lacking information on prevalence
and genomic diversity of hMPVs in Eastern India.
Additional material
Additional file 1: Alignment of the Deduced amino acid sequence
of partial F ORF. Multiple alignment of aa sequences of F protein gene
of hMPV strains from Kolkata. The prototype strain CAN97-83 (GenBank
accession number AY485253) is displayed as consensus sequence.
Identical residues are indicated by dots and dashes represent gaps.
Cysteine residues are marked with asterisks. Potential N-glycosylation
sites are underlined. Cleavage site is boxed.
Abbreviations
aa: Amino acid; ARTI: Acute respiratory tract infection; hMPV: Human
metapneumovirus; nt: Nucleotide; RSV: Respiratory Syncytial virus; RT-PCR:
Reverse transcriptase-polymerase chain reaction.
Acknowledgements
The study was supported by financial assistance from Indian Council of
Medical Research (ICMR), New Delhi. We acknowledge technical assistance
and data entry extended by Malay De Sarkar during the study. A. S. Agrawal
was supported by Senior Research Fellowship from ICMR, Govt. of India.
Authors’ contributions
ASA drafted the manuscript and performed phylogenetic analysis. TR
performed screening of clinical samples by RT-PCR. SG collected clinical
samples and analyzed epidemiological data. MCS conceived the study,
provided guidance and editing of manuscript. All authors read and
approved the final manuscript.
Competing interests
The authors declare that they have no competing interests.
Received: 13 November 2010 Accepted: 12 February 2011
Published: 12 February 2011
References
1. Murray J, Loney C, Murphy LB, Graham S, Yeo RP: Characterization of
monoclonal antibodies raised against recombinant respiratory syncytial virus
nucleocapsid (N) protein: identification of a region in the carboxy terminus
of N involved in the interaction with P protein. Virology 2001, 289:252-261.
2. van den Hoogen BG, de Jong JC, Groen J, Kuiken T, de Groot R,
Fouchier RA, Fouchier RA, Osterhaus AD: A newly discovered human
pneumovirus isolated from young children with respiratory tract disease.
Nat Med 2001, 7:719-724.
3. Peret TC, Boivin G, Li Y, Couillard M, Humphrey C, Osterhaus AD, Erdman DD,
Anderson LJ: Characterization of human metapneumoviruses isolated
from patients in North America. J Infect Dis 2002, 185(11):1660-1663.
4. Stockton J, Stephenson I, Fleming D, Zambon M: Human
metapneumovirus as a cause of community-acquired respiratory illness.
Emerg Infect Dis 2002, 8:897-901.
5. Bastien N, Normand S, Taylor T, Ward D, Peret TC, Boivin G, Anderson LJ,
Li Y: Sequence analysis of the N, P, M and F genes of Canadian human
metapneumovirus strains. Virus Res 2003, 93:51-62.
6. Osterhaus A, Fouchier R: Human metapneumovirus in the community.
Lancet 2003, 361:890-891.
7. van den Hoogen BG, Bestebroer TM, Osterhaus ADME, Fouchier RAM:
Analysis of the genomic sequence of a human metapneumovirus.
Virology 2002, 295:119-132.
8. Hall CB, Walsh EE, Long CE, Schnabel KC: Immunity to and frequency of
reinfection with respiratory syncytial virus. J Infect Dis 1991,
163:693-698.
Agrawal et al. Virology Journal 2011, 8:67
http://www.virologyj.com/content/8/1/67
9.
10.
11.
12.
13.
14.
15.
16.
17.
18.
19.
20.
21.
22.
23.
24.
25.
26.
27.
28.
Levine S, Klaiber-Franco R, Paradiso PR: Demonstration that glycoprotein G
is the attachment protein of respiratory syncytial virus. J Gen Virol 1987,
68:2521-2524.
Biacchesi S, Skiadopoulos MH, Boivin G, Hanson CT, Murphy PR, Collins PL,
Buchholz UJ: Genetic diversity between human metapneumovirus
subgroups. Virology 2003, 315:1-9.
Van Den Hoogen BG, Herfst S, Sprong L, Cane PA, Forleo-Neto E, de
Swart RL, Osterhaus AD, Fouchier RA: Antigenic and genetic variability of
human metapneumoviruses. Emerg Infect Dis 2004, 10:658-666.
Ludewick H, Abed Y, van Niekerk N, Boivin G, Klugman K, Madhi S: Human
metapneumovirus genetic variability, South Africa. Emerg Infect Dis 2005,
11:1074-1078.
Huck B, Scharf G, Neumann-Haefelin D, Weigl J, Falcone V: Novel human
metapneumovirus sublineage. Emerg Infect Dis 2006, 12:147-150.
Gerna G, Campanini G, Rovida F, Sarasini A, Lilleri D, Paolucci S, Marchi A,
Baldanti F, Revello MG: Changing circulation rate of human
metapneumovirus strains and types among hospitalized pediatric
patients during three consecutive winter-spring seasons. Arch Virol 2005,
150:2365-2375.
Agapov E, Sumino KC, Gaudreault-Keener M, Storch GA, Holtzman MJ:
Genetic variability of human metapneumovirus infection: Evidence of a
shift in viral genotype without change in illness. J Infect Dis 2006,
193:396-403.
MacKay IM, Bialasiewicz S, Jacob KC, McQueen E, Arden KE, Nissen MD,
Sloots TP: Genetic diversity of human metapneumovirus over 4
consecutive years in Australia. J Infect Dis 2006, 193:396-403.
Skiadopulos M, Biacchesi S, Buchholz U, Roggs J, Surman S, AmaroCarambotE, McAuliffe J, Elkins W, St Claire M, Collins P, Murphy B: The two
major Human Metapneumovirus genetic lineages are highly related
antigenically, and the Fusion (F) protein is a major contributor to this
antigenic relatedness. J Virol 2004, 78:6927-6937.
Johnson PR, Spriggs MK, Olmsted RA, Collins PL: The G glycoprotein of
human respiratory syncytial viruses of subgroups A and B: Extensive
sequence divergence between antigenically related proteins. Proc Natl
Acad Sci USA 1987, 84:5625-5629.
Ahmad OB, Lopez AD, Inoue M: The decline in child mortality: a
reappraisal. Bull WHO 2000, 78:1175-1191.
Reddaiah VP, Kapoor SK: Acute respiratory infections in rural under fives.
Ind J pediatr 1988, 35:424-426.
Williams BG, Gouws E, Boschi-Pinto C, Bryce J, Dye C: Estimates of worldwide distribution of child deaths from acute respiratory infections.
Lancet Infect Dis 2002, 2:25-32.
Agrawal AS, Sarkar M, Chakrabarti S, Rajendran K, Kaur H, Mishra AC,
Chatterjee MK, Naik TN, Chadha MS, Chawla-Sarkar M: Comparative
evaluation of real-time PCR and conventional RT-PCR during a 2 year
surveillance for influenza and respiratory syncytial virus among children
with acute respiratory infections in Kolkata, India, reveals a distinct
seasonality of infection. J Med Microbiol 2009, 58:1616-1622.
Agrawal AS, Sarkar M, Ghosh S, Roy T, Chakrabarti S, Lal R, Mishra AC,
Chadha MS, Chawla-Sarkar M: Genetic characterization of circulating
seasonal Influenza A viruses (2005-2009) revealed introduction of
oseltamivir resistant H1N1 strains during 2009 in eastern India. Infect
Genet Evol 2010, 10:1188-1198.
Banerjee S, Bharaj P, Sullender W, Kabra SK, Broor S: Human
metapneomovirus infections among children with acute respiratory
infections among children with acute respiratory infections seen in a
large referral hospital in India. J Clin Virol 2007, 38:70-72.
Vabret A, Legrand L, Dina J, Gouarin S, Petitjean-Lecherbonnier J,
Pozzetro B, Ginevra C, Freymuth F: Development of three multiplex RTPCR assays for the detection of 12 respiratory RNA viruses. J Virol
Methods 2005, 126:53-63.
Lund O, Tolstrup N, Gooley AA, Williams KL, Brunak S: NetOglyc: prediction
of mucin type O-glycosylation sites based on sequence context and
surface accessibility. Glycoconjugate J 1998, 15:115-130.
Gupta R, Jung E, Brunak S: Prediction of N-glycosylation sites in human
proteins. NetNGlyc 1.0 2004 [http://www.cbs.dtu.dk/services/NetNGlyc/].
Kumar S, Dudley J, Nei M, Tamura K: MEGA: A biologist-centric software
for evolutionary analysis of DNA and protein sequences. Briefings in
Bioinformatics 2008, 9:299-306.
Page 8 of 8
29. Peret TC, Abed Y, Anderson LJ, Erdman DD, Boivin G: Sequence
polymorphism of the predicted human metapneumovirus G
glycoprotein. J Gen Virol 2004, 85:679-686.
30. Lopez JA, Bustos R, Orvell C, Berois M, Arbiza J, Garcia-Barreno B, Melero JA:
Antigenic structure of human respiratory syncytial virus fusion
glycoprotein. J Virol 1998, 72:6922-6928.
31. Organizacion Panamericana de la Salud (OPS): Infecciones respiratorias
agudas en las Ameracas. Biol Epidemiol 1995, 16:1-5.
32. Boivin G, De Serres G, Côté S, Gilca R, Abed Y, Rochette L, Bergeron MG,
Déry P: Human metapneumovirus infections in hospitalized children.
Emerg Infect Dis 2003, 9(6):634-640.
33. Prins JM, Wolthers KC: Human metapneumovirus: a new pathogen in
children and adults. Neth J Med 2004, 62:177-179.
34. Van den Hoogen BG, van Doornum GJ, Fockens JC, Cornelissen JJ,
Beyer WE, de Groot R, Osterhaus AD, Fouchier RA: Prevalence and clinical
symptoms of human metapneumovirus infection in hospitalized
patients. J Infect Dis 2003, 188:1571-1577.
35. Boivin G, Mackay I, Sloots TP, Madhi S, Freymuth F, Wolf D, Shemer- Avni Y,
Ludewick H, Gray GC, LeBlanc E: Global genetic diversity of human
metapneumovirus fusion gene. Emerg Infect Dis 2004, 10:1154-1157.
36. Falsey AR, Erdman D, Anderson LJ, Walsh EE: Human metapneumovirus
infections in young and elderly adults. J Infect Dis 2003, 187:785-90.
37. Luchsinger V, Escobar C, Avendan˜ o L: Deteccio´n de metapneumovirus
humano en nin˜ os hospitalizados por infeccio´n respiratoria aguda baja
en Santiago, Chile (hMPV en Chile). Rev Me´d Chile 2005, 133:1059-1064.
38. Galiano M, Videla C, Puch SS, Martinez A, Echavarria M, Carballal G:
Evidence of human metapneumovirus in children in Argentina. J Med
Virol 2004, 72:299-303.
39. Freymuth F, Vabret A, Legrand L, Eterradossi N, Lafay-Delaire F, Brouard J,
Guillois B: Presence of the new human metapneumovirus in French
children with bronchiolitis. Pediatr Infect Dis J 2003, 22:92-94.
40. Shek LP, Lee BW: Epidemiology and seasonality of respiratory tract virus
infections in the tropics. Paediatr Respir Rev 2003, 4:105-111.
41. Esper F, Boucher D, Weibel C, Martinello RA, Kahn JS: Human
metapneumovirus infection in the United States: Clinical manifestations
associated with a newly emerging respiratory infection in children.
Pediatrics 2003, 111:1407-1410.
42. Ishiguro N, Ebihara T, Endo R, Ma X, Kikuta H, Ishiko H, Kobayashi K: High
genetic diversity of the attachment (G) protein of human
metapneumovirus. J Clin Microbiol 2004, 42:3406-3414.
43. Galiano M, Trento A, Ver L, Carballal G, Videla C: Genetic Heterogeneity of
G and F Protein Genes From Argentinean Human Metapneumovirus
Strains. J med virol 2006, 78:631-637.
44. Garcia O, Martin M, Dopazo J, Arbiza J, Frabasile S, Russi J, Perez-Brena P,
Martinez I, Garcia-Barreno B, Melero JA: Evolutionary pattern of human
respiratory syncytial virus (subgroup A): Cocirculating lineages and
correlation of genetic and antigenic changes in the G glycoprotein.
J Virol 1994, 68:5448-5459.
45. Peret TC, Hall CB, Schnabel KC, Golub JA, Anderson LJ: Circulation patterns
of genetically distinct group A and B strains of human respiratory
syncytial virus in a community. J Gen Virol 1998, 79(Pt 9):2221-2229.
46. Martinez I, Valdes O, Delfraro A, Arbiza J, Russi J, Melero JA: Evolutionary
pattern of the G glycoprotein of human respiratory syncytial viruses
from antigenic group B: The use of alternative termination codons and
lineage diversification. J Gen Virol 1999, 80:125-130.
doi:10.1186/1743-422X-8-67
Cite this article as: Agrawal et al.: Genetic variability of attachment (G)
and Fusion (F) protein genes of human metapneumovirus strains
circulating during 2006-2009 in Kolkata, Eastern India. Virology Journal
2011 8:67.
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