Characterization of a Large Cluster of Influenza A(H1N1)pdm09 Viruses Cross-Resistant to Oseltamivir and Peramivir during the 2013-2014 Influenza Season in Japan

Abstract

, 2,482 influenza 2009 pandemic A(H1N1) [A(H1N1)pdm09] viruses were screened in Japan for the H275Y substitution in their neuraminidase (NA) protein, which confers cross-resistance to oseltamivir and peramivir. We found that a large cluster of the H275Y mutant virus was present prior to the main influenza season in Sapporo/ Hokkaido, with the detection rate for this mutant virus reaching 29% in this area. Phylogenetic analysis suggested the clonal expansion of a single mutant virus in Sapporo/Hokkaido. To understand the reason for this large cluster, we examined the in vitro and in vivo properties of the mutant virus. We found that it grew well in cell culture, with growth comparable to that of the wildtype virus. The cluster virus also replicated well in the upper respiratory tract of ferrets and was transmitted efficiently between ferrets by way of respiratory droplets. Almost all recently circulating A(H1N1)pdm09 viruses, including the cluster virus, possessed two substitutions in NA, V241I and N369K, which are known to increase replication and transmission fitness. A structural analysis of NA predicted that a third substitution (N386K) in the NA of the cluster virus destabilized the mutant NA structure in the presence of the V241I and N369K substitutions. Our results suggest that the cluster virus retained viral fitness to spread among humans and, accordingly, caused the large cluster in Sapporo/Hokkaido. However, the mutant NA structure was less stable than that of the wild-type virus. Therefore, once the wild-type virus began to circulate in the community, the mutant virus could not compete and faded out. The neuraminidase (NA) inhibitors oseltamivir and zanamivir are recommended by the World Health Organization (WHO) for the treatment of influenza patients, as well as for chemoprophylaxis (1). In Japan, four NA inhibitors, oseltamivir, zanamivir, peramivir, and laninamivir, are approved and prescribed at the highest frequency in the world. Therefore, the surveillance of NA inhibitor-resistant viruses is important for public health authorities and clinicians to control influenza. We have been conducting such surveillance throughout Japan since 1999 (2-6).During the 2006-2007 influenza season, an oseltamivir-resistant former seasonal influenza A(H1N1) virus was first reported in Norway; this virus then spread rapidly and predominated globally until it was replaced by the pandemic A(H1N1) [A(H1N1)pdm09] virus in 2009 (7). The resistant A(H1N1) virus possessed a histidine-to-tyrosine substitution at position 275 (N1 numbering, H275Y) in its NA protein, which was responsible for its drug resistance (7). Four additional NA substitutions, R222Q, V234M, D344N, and D354G, were shown to compensate for the detrimental effect of the H275Y substitution on viral fitness, thereby making the mutant virus more transmissible in the community than the wild-type virus (8-11). However, since the A(H1N1)pdm09 virus began circulating globally in 2009, the H275Y mutant A(H1N1)pdm09 virus has been detected in various regions of t...

Community- and hospital-acquired infections with oseltamivir- and peramivir-resistant influenza A(H1N1)pdm09 viruses during the 2015–2016 season in Japan

“…This incidence is similar to the rate (1.0 %) reported by National Institute of Infectious Disease, Tokyo, Japan during 2015–2016 season (as of 4 March, 2016) [10]. However, this incidence was lower than that reported during the 2013–2014 season (4.1 %) [7]. …”

“…Viruses carrying this mutation possess cross resistance to oseltamivir and peramivir but remain susceptible to zanamivir and laninamivir [2, 3]. Community-acquired oseltamivir- and peramivir-resistant viruses with the H275Y mutation have been recently identified in several countries [5–7]. Community spread of resistant viruses raises concerns regarding their potential to replace sensitive strains as in the case of the resistant seasonal A(H1N1) viruses that spread globally between 2007 and 2008 [2, 3].…”

Characterization of an A (H1N1)pdm09 Virus Imported from India in March 2015

“…Since the A(H1N1)pdm09 virus began circulating globally in 2009, NA inhibitor-resistant viruses have been detected sporadically. However, community clusters of oseltamivir and peramivir cross-resistant viruses occurred in Australia in 2011, and in Japan in 2013-2014, respectively (10,11). Significant numbers of these resistant viruses were also detected in the United States and China during the 2013-2014 season (6,12,13).…”

“…We determined the susceptibility of the Indian-origin A(H1N1)pdm09 virus to currently available NA inhibitors including, oseltamivir, peramivir, zanamivir, and laninamivir by using a fluorescent NA inhibition assay (11). The results are expressed as the drug concentration required to inhibit NA activity by 50z (IC 50 ).…”

Standardizing the influenza neuraminidase inhibition assay among United States public health laboratories conducting virological surveillance

“…In 2012, in efforts to harmonize the interpretation and reporting of IC 50 data, the WHO Expert Working Group for GISRS on Surveillance of Antiviral Susceptibility (WHO-AVWG) agreed on criteria to define influenza viruses as exhibiting normal, reduced (RI) or highly reduced (HRI) NA inhibition, based on the fold change of their IC 50 compared to reference IC 50 values (WHO, 2012). These criteria have been helpful in interpretation and reporting of NI assay data generated by different WHO Collaborating Centers (OkomoAdhiambo et al, 2014;Takashita et al, 2014Takashita et al, , 2015a, and for providing annual global updates (Meijer et al, 2014;Takashita et al, 2015b). Viruses of N1 subtype carrying the H275Y substitution in the NA (H274Y in N2 subtype) and A(H3N2) viruses carrying E119V or R292K substitutions consistently demonstrate HRI by oseltamivir in NI assays (Meijer et al, 2014;Okomo-Adhiambo et al, 2014;Takashita et al, 2015b).…”