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ABSTRACT: Please cite this paper as: Balish et al. (2012) Analytical detection of influenza A(H3N2)v and other A variant viruses from the USA by rapid influenza diagnostic tests. Influenza and Other Respiratory Viruses DOI:10.1111/irv.12017. Background The performance of rapid influenza diagnostic tests (RIDTs) that detect influenza viral nucleoprotein (NP) antigen has been reported to be variable. Recent human infections with variant influenza A viruses that are circulating in pigs prompted the investigation of the analytical reactivity of RIDTs with these variant viruses. Objectives To determine analytical reactivity of seven FDA-cleared RIDTs with influenza A variant viruses in comparison with the reactivity with recently circulating seasonal influenza A viruses. Methods Tenfold serial dilutions of cell culture-grown seasonal and variant influenza A viruses were prepared and tested in duplicate with seven RIDTs. Results All RIDTs evaluated in this study detected the seasonal influenza A(H3N2) virus, although detection limits varied among assays. All but one examined RIDT identified the influenza A(H1N1)pdm09 virus. However, only four of seven RIDTs detected all influenza A(H3N2)v, A(H1N2)v, and A(H1N1)v viruses. Reduced sensitivity of RIDTs to variant influenza viruses may be due to amino acid differences between the NP proteins of seasonal viruses and the NP proteins from viruses circulating in pigs. Conclusions Clinicians should be aware of the limitations of RIDTs to detect influenza A variant viruses. Specimens from patients with influenza-like illness in whom H3N2v is suspected should be sent to public health laboratories for additional diagnostic testing.
Influenza and Other Respiratory Viruses 09/2012; · 4.16 Impact Factor
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Stephen Lindstrom, Rebecca Garten,
Amanda Balish,
Bo Shu,
Shannon Emery,
LaShondra Berman,
Nathelia Barnes,
Katrina Sleeman,
Larisa Gubareva,
Julie Villanueva,
Alexander Klimov
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ABSTRACT: During July-December 2011, a variant virus, influenza A(H3N2)v, caused 12 human cases of influenza. The virus contained genes originating from swine, avian, and human viruses, including the M gene from influenza A(H1N1)pdm09 virus. Influenza A(H3N2)v viruses were antigenically distinct from seasonal influenza viruses and similar to proposed vaccine virus A/Minnesota/11/2010.
Emerging Infectious Diseases 05/2012; 18(5):834-7. · 6.79 Impact Factor
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ABSTRACT: Genetic analysis of pandemic 2009 influenza A (H1N1; H1N1pdm09) virus was undertaken to understand virus evolution during 2009 and 2010 in India. Surveillance of influenza viruses from July 2009 to December 2010 revealed major peaks of circulating H1N1pdm09 viruses in August-September and December-January 2009 and then in August-September 2010. To understand the diversity of the H1N1pdm09 virus, selected specimens (n = 23) from 2009 or 2010 were characterized by nucleotide sequence determination of the HA1 subunit of the HA gene. Phylogenetic analysis revealed that 22 clustered with clade 7 viruses characterized by S203T mutations, whereas one virus from 2010 fell within clade 6. None of the viruses from either 2009 or 2010 formed a monophyletic group, suggesting a continuum of independent introduction of circulating viral strains. Amino acid analysis revealed minor amino acid changes in the antigenic or receptor-binding domains. Importantly, we observed mutations that were also present in 1918 pandemic virus, which includes S183P in 4 and S185T mutation in 3 of 13 viruses analyzed from 2010, while none of the 2009 viruses carried these mutations. Whether antibody-mediated pressure is imposing such changes remains to be determined. Continued genetic surveillance is warranted to monitor pathogenicity as the virus evolves to acquire new features.
Journal of Medical Virology 03/2012; 84(3):386-93. · 2.82 Impact Factor
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Uzma Bashir Aamir,
Nazish Badar,
Muhammad Rashid Mehmood,
Nadia Nisar,
Rana Muhammad Suleman,
Shehzad Shaukat,
Salman Sharif,
Jaleel Kamran,
Syed Sohail Zahoor Zaidi,
Birjees Mazher Kazi,
Larisa Gubareva,
Xiyan Xu, Rebecca Garten,
Alexander Klimov
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ABSTRACT: In early 2009, a novel influenza A(H1N1) virus that emerged in Mexico and United States rapidly disseminated worldwide. The spread of this virus caused considerable morbidity with over 18000 recorded deaths. The new virus was found to be a reassortant containing gene segments from human, avian and swine influenza viruses.
The first case of human infection with A(H1N1)pdm09 in Pakistan was detected on 18(th) June 2009. Since then, 262 laboratory-confirmed cases have been detected during various outbreaks with 29 deaths (as of 31(st) August 2010). The peak of the epidemic was observed in December with over 51% of total respiratory cases positive for influenza. Representative isolates from Pakistan viruses were sequenced and analyzed antigenically. Sequence analysis of genes coding for surface glycoproteins HA and NA showed high degree of high levels of sequence identity with corresponding genes of regional viruses circulating South East Asia. All tested viruses were sensitive to Oseltamivir in the Neuraminidase Inhibition assays.
Influenza A(H1N1)pdm09 viruses from Pakistan form a homogenous group of viruses. Their HA genes belong to clade 7 and show antigenic profile similar to the vaccine strain A/California/07/2009. These isolates do not show any amino acid changes indicative of high pathogenicity and virulence. It is imperative to continue monitoring of these viruses for identification of potential variants of high virulence or drug resistance.
PLoS ONE 01/2012; 7(8):e41866. · 4.09 Impact Factor
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Bo Shu, Rebecca Garten,
Shannon Emery,
Amanda Balish,
Lynn Cooper,
Wendy Sessions,
Varough Deyde,
Catherine Smith,
LaShondra Berman,
Alexander Klimov,
Stephen Lindstrom,
Xiyan Xu
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ABSTRACT: Swine influenza viruses (SIV) have been recognized as important pathogens for pigs and occasional human infections with swine origin influenza viruses (SOIV) have been reported. Between 1990 and 2010, a total of twenty seven human cases of SOIV infections have been identified in the United States. Six viruses isolated from 1990 to 1995 were recognized as classical SOIV (cSOIV) A(H1N1). After 1998, twenty-one SOIV recovered from human cases were characterized as triple reassortant (tr_SOIV) inheriting genes from classical swine, avian and human influenza viruses. Of those twenty-one tr_SOIV, thirteen were of A(H1N1), one of A(H1N2), and seven of A(H3N2) subtype. SOIV characterized were antigenically and genetically closely related to the subtypes of influenza viruses circulating in pigs but distinct from contemporary influenza viruses circulating in humans. The diversity of subtypes and genetic lineages in SOIV cases highlights the importance of continued surveillance at the animal-human interface.
Virology 11/2011; 422(1):151-60. · 3.35 Impact Factor
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Parvaiz A Koul,
Muneer A Mir,
Nargis K Bali,
Mamta Chawla-Sarkar,
Mehuli Sarkar,
Samander Kaushik,
U H Khan,
Feroze Ahmad, Rebecca Garten,
Renu B Lal,
Shobha Broor
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ABSTRACT: With the emergence of pandemic influenza A (2009A/H1N1) virus in India, we sought to determine the prevalence and clinical presentations of seasonal and pandemic influenza viruses among acute respiratory illness (ARI) patients from Srinagar, a temperate climate area in northern India, during the peak winter season.
Combined throat and nasal swabs, obtained from 194 (108 male) presenting with ARI from January to March 2010 (Week 53-week 10), were tested by RT-PCR for influenza A and B, including 2009A/H1N1 viruses. HA1 gene of selected 2009A/H1N1-positive samples was sequenced, and phylogenetic analysis was carried out.
Twenty-one (10·8%, age 15-80 years, median age 40 years) patients tested positive for influenza viruses: 13 (62%) for 2009A/H1N1 virus, 6 (28·5%) for seasonal influenza A (H3N2), and 2 (9·5%) for influenza B. Twelve of the 13 patients with 2009A/H1N1 presented with febrile ARI, and eight had associated comorbidities. All of the patients recovered. Phylogenetic analysis of HA gene (n = 8) revealed that all strains from Srinagar clustered in 2009A/H1N1 clade seven along with the other 2009A/H1N1 strains from India. Amino acid substitutions in the HA protein defining clade seven (P83S, S203T, and I321V) were found in almost all isolates from Srinagar.
Both seasonal and 2009A/H1N1 viruses appear to be associated with ARI in Srinagar. The 2009A/H1N1 in Srinagar is genetically similar to globally circulating clade 7 strains, with unique signature sequences in the HA gene. Further investigations into ascertain the role of these mutations in possible alteration of the virulence and transmissibility of the virus are needed.
Influenza and Other Respiratory Viruses 05/2011; 5(6):e521-7. · 4.16 Impact Factor
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Li-Mei Chen,
Pierre Rivailler,
Jaber Hossain,
Paul Carney,
Amanda Balish,
Ijeoma Perry,
C Todd Davis, Rebecca Garten,
Bo Shu,
Xiyan Xu,
Alexander Klimov,
James C Paulson,
Nancy J Cox,
Sabrina Swenson,
James Stevens,
Amy Vincent,
Marie Gramer,
Ruben O Donis
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ABSTRACT: The evolution of classical swine influenza viruses receptor specificity preceding the emergence of the 2009 H1N1 pandemic virus was analyzed in glycan microarrays. Classical swine influenza viruses from the α, β, and γ antigenic clusters isolated between 1945 and 2009 revealed a binding profile very similar to that of 2009 pandemic H1N1 viruses, with selectivity for α2-6-linked sialosides and very limited binding to α2-3 sialosides. Despite considerable genetic divergence, the 'human-like' H1N1 viruses circulating in swine retained strong binding preference for α2-6 sialylated glycans. Interspecies transmission of H1N1 influenza viruses from swine to humans or from humans to swine has not driven selection of viruses with distinct novel receptor binding specificities. Classical swine and human seasonal H1N1 influenza viruses have conserved specificity for similar α2-6-sialoside receptors in spite of long term circulation in separate hosts, suggesting that humans and swine impose analogous selection pressures on the evolution of receptor binding function.
Virology 02/2011; 412(2):401-10. · 3.35 Impact Factor
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Kulbhushan Sharma,
Shashank Tripathi,
Priya Ranjan,
Purnima Kumar, Rebecca Garten,
Varough Deyde,
Jacqueline M Katz,
Nancy J Cox,
Renu B Lal,
Suryaprakash Sambhara,
Sunil K Lal
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ABSTRACT: Double-stranded RNA dependent protein kinase (PKR) is a key regulator of the anti-viral innate immune response in mammalian cells. PKR activity is regulated by a 58 kilo Dalton cellular inhibitor (P58(IPK)), which is present in inactive state as a complex with Hsp40 under normal conditions. In case of influenza A virus (IAV) infection, P58(IPK) is known to dissociate from Hsp40 and inhibit PKR activation. However the influenza virus component responsible for PKR inhibition through P58(IPK) activation was hitherto unknown.
Human heat shock 40 protein (Hsp40) was identified as an interacting partner of Influenza A virus nucleoprotein (IAV NP) using a yeast two-hybrid screen. This interaction was confirmed by co-immunoprecipitation studies from mammalian cells transfected with IAV NP expressing plasmid. Further, the IAV NP-Hsp40 interaction was validated in mammalian cells infected with various seasonal and pandemic strains of influenza viruses. Cellular localization studies showed that NP and Hsp40 co-localize primarily in the nucleus. During IAV infection in mammalian cells, expression of NP coincided with the dissociation of P58(IPK) from Hsp40 and decrease PKR phosphorylation. We observed that, plasmid based expression of NP in mammalian cells leads to decrease in PKR phosphorylation. Furthermore, inhibition of NP expression during influenza virus replication led to PKR activation and concomitant increase in eIF2α phosphorylation. Inhibition of NP expression also led to reduced IRF3 phosphorylation, enhanced IFN β production and concomitant reduction of virus replication. Taken together our data suggest that NP is the viral factor responsible for P58(IPK) activation and subsequent inhibition of PKR-mediated host response during IAV infection.
Our findings demonstrate a novel role of IAV NP in inhibiting PKR-mediated anti-viral host response and help us understand P58(IPK) mediated inhibition of PKR activity during IAV infection.
PLoS ONE 01/2011; 6(6):e20215. · 4.09 Impact Factor
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Pauline Terebuh,
Christopher W Olsen,
Jennifer Wright,
Alexander Klimov,
Alexander Karasin,
Karla Todd,
Hong Zhou,
Henrietta Hall,
Xiyan Xu,
Tim Kniffen,
David Madsen, Rebecca Garten,
Carolyn B Bridges
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ABSTRACT: Triple-reassortant (tr) viruses of human, avian, and swine origin, including H1N1, H1N2, and H3N2 subtypes, emerged in North American swine herds in 1998 and have become predominant. While sporadic human infections with classical influenza A (H1N1) and with tr-swine influenza viruses have been reported, relatively few have been documented in occupationally exposed swine workers (SW).
We conducted a 2-year (2002-2004) prospective cohort study of transmission of influenza viruses between pigs and SW from a single pork production company in Iowa. Respiratory samples were collected and tested for influenza viruses from SW and from pigs under their care through surveillance for influenza-like illnesses (ILI). Serial blood samples from study participants were tested by hemagglutination inhibition (HI) for antibody seroconversion against human and swine influenza viruses (SIV), and antibody seroprevalence was compared to age-matched urban Iowa blood donors.
During the first year, 15 of 88 SW had ILI and were sampled; all were culture-negative for influenza. During the second year, 11 of 76 SW had ILI and were sampled; one was culture-positive for a human seasonal H3N2 virus. Among 20 swine herd ILI outbreaks sampled, influenza A virus was detected by rRT-PCR from 17 with 11 trH1N1 and five trH3N2 virus isolates cultured. During both years, HI geometric mean titers were significantly higher among SW compared to blood donor controls for three SIV: classical swine Sw/WI/238/97 (H1N1), tr Sw/IN/9K035/99 (H1N2), and trSw/IA/H02NJ56371/02 (H1N1)] (P < 0·0001).
SW had serologic evidence for infection with both swine and human influenza viruses and were exposed to diverse influenza virus strains circulating in pigs. Influenza virus surveillance among pigs and SW should be encouraged to better understand cross-species transmission and diversity of influenza viruses at the human-swine interface.
Influenza and Other Respiratory Viruses 11/2010; 4(6):387-96. · 4.16 Impact Factor
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Pauline Terebuh,
Christopher W. Olsen,
Jennifer Wright,
Alexander Klimov,
Alexander Karasin,
Karla Todd,
Hong Zhou,
Henrietta Hall,
Xiyan Xu,
Tim Kniffen,
David Madsen, Rebecca Garten,
Carolyn B. Bridges
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ABSTRACT: Please cite this paper as: Terebuh et al. (2010) Transmission of influenza A viruses between pigs and people, Iowa, 2002–2004. Influenza and Other Respiratory Viruses 4(6), 387–396.Background Triple-reassortant (tr) viruses of human, avian, and swine origin, including H1N1, H1N2, and H3N2 subtypes, emerged in North American swine herds in 1998 and have become predominant. While sporadic human infections with classical influenza A (H1N1) and with tr-swine influenza viruses have been reported, relatively few have been documented in occupationally exposed swine workers (SW).Methods We conducted a 2-year (2002–2004) prospective cohort study of transmission of influenza viruses between pigs and SW from a single pork production company in Iowa. Respiratory samples were collected and tested for influenza viruses from SW and from pigs under their care through surveillance for influenza-like illnesses (ILI). Serial blood samples from study participants were tested by hemagglutination inhibition (HI) for antibody seroconversion against human and swine influenza viruses (SIV), and antibody seroprevalence was compared to age-matched urban Iowa blood donors.Results During the first year, 15 of 88 SW had ILI and were sampled; all were culture-negative for influenza. During the second year, 11 of 76 SW had ILI and were sampled; one was culture-positive for a human seasonal H3N2 virus. Among 20 swine herd ILI outbreaks sampled, influenza A virus was detected by rRT-PCR from 17 with 11 trH1N1 and five trH3N2 virus isolates cultured. During both years, HI geometric mean titers were significantly higher among SW compared to blood donor controls for three SIV: classical swine Sw/WI/238/97 (H1N1), tr Sw/IN/9K035/99 (H1N2), and trSw/IA/H02NJ56371/02 (H1N1)] (P < 0·0001).Conclusions SW had serologic evidence for infection with both swine and human influenza viruses and were exposed to diverse influenza virus strains circulating in pigs. Influenza virus surveillance among pigs and SW should be encouraged to better understand cross-species transmission and diversity of influenza viruses at the human–swine interface.
Influenza and Other Respiratory Viruses 10/2010; 4(6):387 - 396. · 4.16 Impact Factor
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James Stevens,
Li-Mei Chen,
Paul J Carney, Rebecca Garten,
Angie Foust,
Jianhua Le,
Barbara A Pokorny,
Ramanunninair Manojkumar,
Jeanmarie Silverman,
Rene Devis,
Karen Rhea,
Xiyan Xu,
Doris J Bucher,
James C Paulson,
James Paulson,
Nancy J Cox,
Alexander Klimov,
Ruben O Donis
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ABSTRACT: Isolation of human subtype H3N2 influenza viruses in embryonated chicken eggs yields viruses with amino acid substitutions in the hemagglutinin (HA) that often affect binding to sialic acid receptors. We used a glycan array approach to analyze the repertoire of sialylated glycans recognized by viruses from the same clinical specimen isolated in eggs or cell cultures. The binding profiles of whole virions to 85 sialoglycans on the microarray allowed the categorization of cell isolates into two groups. Group 1 cell isolates displayed binding to a restricted set of alpha2-6 and alpha2-3 sialoglycans, whereas group 2 cell isolates revealed receptor specificity broader than that of their egg counterparts. Egg isolates from group 1 showed binding specificities similar to those of cell isolates, whereas group 2 egg isolates showed a significantly reduced binding to alpha2-6- and alpha2-3-type receptors but retained substantial binding to specific O- and N-linked alpha2-3 glycans, including alpha2-3GalNAc and fucosylated alpha2-3 glycans (including sialyl Lewis x), both of which may be important receptors for H3N2 virus replication in eggs. These results revealed an unexpected diversity in receptor binding specificities among recent H3N2 viruses, with distinct patterns of amino acid substitution in the HA occurring upon isolation and/or propagation in eggs. These findings also suggest that clinical specimens containing viruses with group 1-like receptor binding profiles would be less prone to undergoing receptor binding or antigenic changes upon isolation in eggs. Screening cell isolates for appropriate receptor binding properties might help focus efforts to isolate the most suitable viruses in eggs for production of antigenically well-matched influenza vaccines.
Journal of Virology 08/2010; 84(16):8287-99. · 5.40 Impact Factor
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ABSTRACT: The M2 blockers amantadine and rimantadine and the neuraminidase (NA) inhibitors (NAIs) oseltamivir and zanamivir are approved by the FDA for use for the control of influenza A virus infections. The 2009 pandemic influenza A (H1N1) viruses (H1N1pdm) are reassortants that acquired M and NA gene segments from a Eurasian adamantane-resistant swine influenza virus. NAI resistance in the H1N1pdm viruses has been rare, and its occurrence is mainly limited to oseltamivir-exposed patients. The pyrosequencing assay has been proven to be a useful tool in surveillance for drug resistance in seasonal influenza A viruses. We provide a protocol which allows the detection of adamantane resistance markers as well as the I43T change, which is unique to the H1N1pdm M2 protein. The protocol also allows the detection of changes at residues V116, I117, E119, Q136, K150, D151, D199, I223, H275, and N295 in the NA, known to alter NAI drug susceptibility. We report on the detection of the first cases of the oseltamivir resistance-conferring mutation H275Y and the I223V change in viruses from the United States using the approach described in this study. Moreover, the assay permits the quick identification of the major NA group (V106/N248, I106/D248, or I106/N248) to which a pandemic virus belongs. Pyrosequencing is well suited for the detection of drug resistance markers and signature mutations in the M and NA gene segments of the pandemic H1N1 influenza viruses.
Antimicrobial Agents and Chemotherapy 12/2009; 54(3):1102-10. · 4.84 Impact Factor
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ABSTRACT: Adamantanes resistance in H3N2 viruses has been increasing since 2000, and in 2005-2006 reached nearly 100% in most countries, with the circulation of the N-lineage. In 2006-2007, however, a significant decrease in resistance was observed in many regions.
To explore potential links between adamantane resistance and the A(H3N2) viruses that circulated between 2006 and 2008.
A total of 1451 Influenza A (H3N2) viruses collected globally in 2001-2008 were screened for the presence of adamantane resistance markers. A subset of 100 viruses representing the broad genetic and geographic spectrum of these viruses was selected for complete genome sequencing and phylogenetic analyses.
Full genome sequence analysis of 2006-2007 viruses revealed co-circulation of four distinct genotypes, designated A-D. Phylogenetic analyses demonstrated reassortment between viruses from the N-lineage and other viruses that had circulated in prior seasons, including those bearing an adamantane sensitive marker. Genotype D viruses became dominant in late 2006-2007 and continued to be the main H3N2 genotype in 2007-2008. Viruses of this genotype retained all N-lineage genome segments except PB2 and NP, which were acquired through reassortment.
The decrease in adamantane resistance at that time was due to transient co-circulation of genotypes that emerged through reassortment. Our findings emphasize the importance of complete genome sequencing in understanding the complex nature of the relationship between influenza virus evolution and antiviral resistance. The recent emergence of the pandemic multi-reassortant H1N1 virus underscores the importance of whole genome sequence monitoring for rapid detection of such unusual and novel strains.
Influenza and Other Respiratory Viruses 11/2009; 3(6):297-314. · 4.16 Impact Factor
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ABSTRACT: The surveillance of seasonal influenza virus susceptibility to neuraminidase (NA) inhibitors was conducted using an NA inhibition assay. The 50% inhibitory concentration values (IC(50)s) of 4,570 viruses collected globally from October 2004 to March 2008 were determined. Based on mean IC(50)s, A(H3N2) viruses (0.44 nM) were more sensitive to oseltamivir than A(H1N1) viruses (0.91 nM). The opposite trend was observed with zanamivir: 1.06 nM for A(H1N1) and 2.54 nM for A(H3N2). Influenza B viruses exhibited the least susceptibility to oseltamivir (3.42 nM) and to zanamivir (3.87 nM). To identify potentially resistant viruses (outliers), a threshold of a mean IC(50) value + 3 standard deviations was defined for type/subtype and drug. Sequence analysis of outliers was performed to identify NA changes that might be associated with reduced susceptibility. Molecular markers of oseltamivir resistance were found in six A(H1N1) viruses (H274Y) and one A(H3N2) virus (E119V) collected between 2004 and 2007. Some outliers contained previously reported mutations (e.g., I222T in the B viruses), while other mutations [e.g., R371K and H274Y in B viruses and H274N in A(H3N2) viruses) were novel. The R371K B virus outlier exhibited high levels of resistance to both inhibitors (>100 nM). A substantial variance at residue D151 was observed among A(H3N2) zanamivir-resistant outliers. The clinical relevance of newly identified NA mutations is unknown. A rise in the incidence of oseltamivir resistance in A(H1N1) viruses carrying the H274Y mutation was detected in the United States and in other countries in the ongoing 2007 to 2008 season. As of March 2008, the frequency of resistance among A(H1N1) viruses in the United States was 8.6% (50/579 isolates). The recent increase in oseltamivir resistance among A(H1N1) viruses isolated from untreated patients raises public health concerns and necessitates close monitoring of resistance to NA inhibitors.
Antimicrobial Agents and Chemotherapy 07/2008; 52(9):3284-92. · 4.84 Impact Factor
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ABSTRACT: Influenza A viruses are hosted by numerous avian and mammalian species, which have shaped their evolution into distinct lineages worldwide. The viral genome consists of eight RNA segments that are frequently exchanged between different viruses via a process known as genetic reassortment. A complete genotype nomenclature is essential to describe gene segment reassortment. Specialized bioinformatic tools to analyze reassortment are not available, which hampers progress in understanding its role in host range, virulence and transmissibility of influenza viruses. To meet this need, we have developed a nomenclature to name influenza A genotypes and implemented a web server, FluGenome (http://www.flugenome.org/), for the assignment of lineages and genotypes. FluGenome provides functions for the user to interrogate the database in different modalities and get detailed reports on lineages and genotypes. These features make FluGenome unique in its ability to automatically detect genotype differences attributable to reassortment events in influenza A virus evolution.
Nucleic Acids Research 08/2007; 35(Web Server issue):W275-9. · 8.03 Impact Factor
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ABSTRACT: A highly discriminative and information-rich diagnostic assay for H5N1 avian influenza would meet immediate patient care needs and provide valuable information for public health interventions, e.g., tracking of new and more dangerous variants by geographic area as well as avian-to-human or human-to-human transmission. In the present study, we have designed a rapid assay based on multilocus nucleic acid sequencing that focuses on the biologically significant regions of the H5N1 hemagglutinin gene. This allows the prediction of viral strain, clade, receptor binding properties, low- or high-pathogenicity cleavage site and glycosylation status. H5 HA genes were selected from nine known high-pathogenicity avian influenza subtype H5N1 viruses, based on their diversity in biologically significant regions of hemagglutinin and/or their ability to cause infection in humans. We devised a consensus pre-programmed pyrosequencing strategy, which may be used as a faster, more accurate alternative to de novo sequencing. The available data suggest that the assay described here is a reliable, rapid, information-rich and cost-effective approach for definitive diagnosis of H5N1 avian influenza. Knowledge of the predicted functional sequences of the HA will enhance H5N1 avian influenza surveillance efforts.
PLoS ONE 02/2006; 1:e95. · 4.09 Impact Factor
