A Bayesian approach to incorporate structural data into the mapping of genotype to antigenic phenotype of influenza A(H3N2) viruses.

Surface antigens of pathogens are commonly targeted by vaccine-elicited antibodies but antigenic variability, notably in RNA viruses such as influenza, HIV and SARS-CoV-2, pose challenges for control by vaccination. For example, influenza A(H3N2) entered the human population in 1968 causing a pandemic and has since been monitored, along with other seasonal influenza viruses, for the emergence of antigenic drift variants through intensive global surveillance and laboratory characterisation. Statistical models of the relationship between genetic differences among viruses and their antigenic similarity provide useful information to inform vaccine development, though accurate identification of causative mutations is complicated by highly correlated genetic signals that arise due to the evolutionary process. Here, using a sparse hierarchical Bayesian analogue of an experimentally validated model for integrating genetic and antigenic data, we identify the genetic changes in influenza A(H3N2) virus that underpin antigenic drift. We show that incorporating protein structural data into variable selection helps resolve ambiguities arising due to correlated signals, with the proportion of variables representing haemagglutinin positions decisively included, or excluded, increased from 59.8% to 72.4%. The accuracy of variable selection judged by proximity to experimentally determined antigenic sites was improved simultaneously. Structure-guided variable selection thus improves confidence in the identification of genetic explanations of antigenic variation and we also show that prioritising the identification of causative mutations is not detrimental to the predictive capability of the analysis. Indeed, incorporating structural information into variable selection resulted in a model that could more accurately predict antigenic assay titres for phenotypically-uncharacterised virus from genetic sequence. Combined, these analyses have the potential to inform choices of reference viruses, the targeting of laboratory assays, and predictions of the evolutionary success of different genotypes, and can therefore be used to inform vaccine selection processes.

Low prevalence of influenza viruses and predominance of A(H3N2) virus with respect to SARS-CoV-2 during the 2021-2022 season in Bulgaria.

Social distancing, mask-wearing, and travel restrictions during the COVID-19 pandemic have significantly impacted the spread of influenza viruses. The objectives of this study were to analyze the pattern of influenza virus circulation with respect to that of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in Bulgaria during the 2021-2022 season and to perform a phylogenetic/molecular analysis of the hemagglutinin (HA) and neuraminidase (NA) sequences of representative influenza strains. Influenza infection was confirmed using real-time reverse transcription polymerase chain reaction in 93 (4.2%) of the 2193 patients with acute respiratory illness tested wherein all detected viruses were subtyped as A(H3N2). SARS-CoV-2 was identified in 377 (24.3%) of the 1552 patients tested. Significant differences in the incidence of influenza viruses and SARS-CoV-2 were found between individual age groups, outpatients/inpatients, and in the seasonal distribution of cases. Two cases of coinfections were identified. In hospitalized patients, the Ct values of influenza viruses at admission were lower in adults aged ≥65 years (indicating higher viral load) than in children aged 0-14 years (p < 0.05). In SARS-CoV-2-positive inpatients, this association was not statistically significant. HA genes of all A(H3N2) viruses analyzed belonged to subclade 3C.2a1b.2a. The sequenced viruses carried 11 substitutions in HA and 5 in NA, in comparison to the vaccine virus A/Cambodia/e0826360/2020, including several substitutions in the HA antigenic sites B and C. This study revealed extensive changes in the typical epidemiology of influenza infection, including a dramatic reduction in the number of cases, diminished genetic diversity of circulating viruses, changes in age, and seasonal distribution of cases.

Emerging triple-reassortant influenza C virus with household-associated infection during an influenza A(H3N2) outbreak, China, 2022.

Recent research have shown that influenza C virus (ICV) has a possible higher clinical impact than previously thought. But knowledge about ICV is limited compared with influenza A and B viruses, due to poor systematic surveillance and inability to propagate. Herein, a case infected with triple reassortant ICV was identified during an influenza A(H3N2) outbreak, which was the first report of ICV infection in mainland China. Phylogenetic analysis showed that this ICV was triple reassortant. Serological evidence revealed that the index case might be related to family-clustering infection. Therefore, it is essential to heighten surveillance for the prevalence and variation of ICV in China, during the COVID-19 pandemic.

Whole-Genome Analysis of Influenza A(H3N2) and B/Victoria Viruses Detected in Myanmar during the COVID-19 Pandemic in 2021.

An influenza circulation was observed in Myanmar between October and November in 2021. Patients with symptoms of influenza-like illness were screened using rapid diagnostic test (RDT) kits, and 147/414 (35.5%) upper respiratory tract specimens presented positive results. All RDT-positive samples were screened by a commercial multiplex real-time polymerase chain reaction (RT-PCR) assay, and 30 samples positive for influenza A(H3N2) or B underwent further typing/subtyping for cycle threshold (Ct) value determination based on cycling probe RT-PCR. The majority of subtyped samples (n = 13) were influenza A(H3N2), while only three were B/Victoria. Clinical samples with low Ct values obtained by RT-PCR were used for whole-genome sequencing via next-generation sequencing technology. All collected viruses were distinct from the Southern Hemisphere vaccine strains of the corresponding season but matched with vaccines of the following season. Influenza A(H3N2) strains from Myanmar belonged to clade 2a.3 and shared the highest genetic proximity with Bahraini strains. B/Victoria viruses belonged to clade V1A.3a.2 and were genetically similar to Bangladeshi strains. This study highlights the importance of performing influenza virus surveillance with genetic characterization of the influenza virus in Myanmar, to contribute to global influenza surveillance during the COVID-19 pandemic.

On the lookout for influenza viruses in Italy during the 2021-2022 season: Along came A(H3N2) viruses with a new phylogenetic makeup of their hemagglutinin.

AIMS: To assess influenza viruses (IVs) circulation and to evaluate A(H3N2) molecular evolution during the 2021-2022 season in Italy. MATERIALS AND METHODS: 12,393 respiratory specimens (nasopharyngeal swabs or broncho-alveolar lavages) collected from in/outpatients with influenza illness in the period spanning from January 1, 2022 (week 2022-01) to May 31, 2022 (week 2022-22) were analysed to identify IV genome and were molecularly characterized by 12 laboratories throughout Italy. A(H3N2) evolution was studied by conducting an in-depth phylogenetic analysis of the hemagglutinin (HA) gene sequences. The predicted vaccine efficacy (pVE) of vaccine strain against circulating A(H3N2) viruses was estimated using the sequence-based Pepitope model. RESULTS: The overall IV-positive rate was 7.2% (894/12,393), all were type A IVs. Almost all influenza A viruses (846/894; 94.6%) were H3N2 that circulated in Italy with a clear epidemic trend, with 10% positivity rate threshold crossed for six consecutive weeks from week 2022-11 to week 2022-16. According to the phylogenetic analysis of a subset of A(H3N2) strains (n=161), the study HA sequences were distributed into five different genetic clusters, all of them belonging to the clade 3C.2a, sub-clade 3C.2a1 and the genetic subgroup 3C.2a1b.2a.2. The selective pressure analysis of A(H3N2) sequences showed evidence of diversifying selection particularly in the amino acid position 156. The comparison between the predicted amino acid sequence of the 2021-2022 vaccine strain (A/Cambodia/e0826360/2020) and the study strains revealed 65 mutations in 59 HA amino acid positions, including the substitution H156S and Y159N in antigenic site B, within major antigenic sites adjacent to the receptor-binding site, suggesting the presence of drifted strains. According to the sequence-based Pepitope model, antigenic site B was the dominant antigenic site and the p(VE) against circulating A(H3N2) viruses was estimated to be -28.9%. DISCUSSION AND CONCLUSION: After a long period of very low IV activity since public health control measures have been introduced to face COVID-19 pandemic, along came A(H3N2) with a new phylogenetic makeup. Although the delayed 2021-2022 influenza season in Italy was characterized by a significant reduction of the width of the epidemic curve and in the intensity of the influenza activity compared to historical data, a marked genetic diversity of the HA of circulating A(H3N2) strains was observed. The identification of the H156S and Y159N substitutions within the main antigenic sites of most HA sequences also suggested the circulation of drifted variants with respect to the 2021-2022 vaccine strain. Molecular surveillance plays a critical role in the influenza surveillance architecture and it has to be strengthened also at local level to timely assess vaccine effectiveness and detect novel strains with potential impact on public health.

Identification and genome sequencing of an influenza H3N2 variant in wastewater from elementary schools during a surge of influenza A cases in Las Vegas, Nevada.

Real-time surveillance of infectious diseases at schools or in communities is often hampered by delays in reporting due to resource limitations and infrastructure issues. By incorporating quantitative PCR and genome sequencing, wastewater surveillance has been an effective complement to public health surveillance at the community and building-scale for pathogens such as poliovirus, SARS-CoV-2, and even the monkeypox virus. In this study, we asked whether wastewater surveillance programs at elementary schools could be leveraged to detect RNA from influenza viruses shed in wastewater. We monitored for influenza A and B viral RNA in wastewater from six elementary schools from January to May 2022. Quantitative PCR led to the identification of influenza A viral RNA at three schools, which coincided with the lifting of COVID-19 restrictions and a surge in influenza A infections in Las Vegas, Nevada, USA. We performed genome sequencing of wastewater RNA, leading to the identification of a 2021-2022 vaccine-resistant influenza A (H3N2) 3C.2a1b.2a.2 subclade. We next tested wastewater samples from a treatment plant that serviced the elementary schools, but we were unable to detect the presence of influenza A/B RNA. Together, our results demonstrate the utility of near-source wastewater surveillance for the detection of local influenza transmission in schools, which has the potential to be investigated further with paired school-level influenza incidence data.

Influenza vaccine effectiveness against influenza A subtypes in Europe: Results from the 2021-2022 I-MOVE primary care multicentre study.

BACKGROUND: In 2021-2022, influenza A viruses dominated in Europe. The I-MOVE primary care network conducted a multicentre test-negative study to measure influenza vaccine effectiveness (VE). METHODS: Primary care practitioners collected information on patients presenting with acute respiratory infection. Cases were influenza A(H3N2) or A(H1N1)pdm09 RT-PCR positive, and controls were influenza virus negative. We calculated VE using logistic regression, adjusting for study site, age, sex, onset date, and presence of chronic conditions. RESULTS: Between week 40 2021 and week 20 2022, we included over 11 000 patients of whom 253 and 1595 were positive for influenza A(H1N1)pdm09 and A(H3N2), respectively. Overall VE against influenza A(H1N1)pdm09 was 75% (95% CI: 43-89) and 81% (95% CI: 45-93) among those aged 15-64 years. Overall VE against influenza A(H3N2) was 29% (95% CI: 12-42) and 25% (95% CI: -41 to 61), 33% (95% CI: 14-49), and 26% (95% CI: -22 to 55) among those aged 0-14, 15-64, and over 65 years, respectively. The A(H3N2) VE among the influenza vaccination target group was 20% (95% CI: -6 to 39). All 53 sequenced A(H1N1)pdm09 viruses belonged to clade 6B.1A.5a.1. Among 410 sequenced influenza A(H3N2) viruses, all but eight belonged to clade 3C.2a1b.2a.2. DISCUSSION: Despite antigenic mismatch between vaccine and circulating strains for influenza A(H3N2) and A(H1N1)pdm09, 2021-2022 VE estimates against circulating influenza A(H1N1)pdm09 were the highest within the I-MOVE network since the 2009 influenza pandemic. VE against A(H3N2) was lower than A(H1N1)pdm09, but at least one in five individuals vaccinated against influenza were protected against presentation to primary care with laboratory-confirmed influenza.

An integrated dual-layer microfluidic platform for multiple respiratory viruses screening.

Currently, the coronavirus disease 2019 (COVID-19) caused by the outbreak of a novel coronavirus (SARS-CoV-2) is spreading rapidly worldwide. Due to the high incidence of influenza coinciding with SARS-CoV-2, rapid detection is crucial to prevent spreading. Here, we present an integrated dual-layer microfluidic platform for specific and highly sensitive SARS-CoV-2, influenza viruses A (FluA) H1N1, H3N2, and influenza virus B (FluB) simultaneous detection. The platform includes a dual microchip (Dµchip) and a portable detection device for real-time fluorescence detection, temperature control and online communication. The Reverse Transcription Loop-mediated Isothermal Amplification (RT-LAMP) and Cas12a cleavage were performed on the Dµchip. The limit of detection (LoD) of the Dµchip assay was 10 copies for SARS-CoV-2, FluA H1N1, H3N2, and FluB RNAs. The Dµchip assay yielded no cross-reactivity against other coronaviruses, so it was suitable for the screening of multiple viruses. Moreover, the positive percentage agreement (PPA) and negative percentage agreement (NPA) of the assay were 97.9% and 100%, respectively, in 75 clinical samples compared to data from RT-PCR-based assays. Furthermore, the assay allowed the detection SARS-CoV-2 and influenza viruses in spiked samples. Overall, the present platform would provide a rapid method for the screening of multiple viruses in hospital emergency, community and primary care settings and facilitate the remote diagnosis and outbreak control of the COVID-19.

What's Next for Flu? Out-of-Season Circulation of Influenza Viruses in Southern Italy, August 2022.

The COVID-19 pandemic has modified the seasonal pattern of respiratory infections. The objective of the present study is to characterize the out-of-season circulation of influenza viruses and an influenza outbreak that occurred in southern Italy in August 2022. Nasopharyngeal swabs collected from patients with influenza-like illnesses (ILI) were tested for the presence of influenza and other respiratory viruses. Epidemiological investigations on 85 patients involved in an influenza outbreak were performed. Sequencing and phylogenetic analysis of hemagglutinin genes was undertaken on samples positive for influenza A. In August 2022, in the Apulia region (Italy), influenza A infection was diagnosed in 19 patients, 18 infected with A/H3N2 and one with A/H1N1pdm09 virus. Seven influenza-positive patients were hospitalized with ILI. A further 17 symptomatic subjects, associated with an influenza outbreak, were also tested; 11 were positive for influenza A/H3N2 virus. Phylogenetic analysis of 12 of the A/H3N2 sequences showed that they all belonged to subclade 3C.2a1b.2a.2. The A/H1N1pdm09 strain belonged to subclade 6B.1A.5a.2. The out-of-season circulation of the influenza virus during the summer months could be linked to changing dynamics in the post-COVID-19 era, as well as to the impact of climate change. Year-round surveillance of respiratory viruses is needed to monitor this phenomenon and to provide effective prevention strategies.

Matrix-Encoding Gene Diversity of 624 Influenza A/H3N2 Genomes Does Not Show Association with Impaired Viral Detection by Commercialized qPCR Assays.

As for the case of SARS-CoV-2, genome sequencing of influenza viruses is of potential interest to raise and address virological issues. Recently, false-negativity of real-time reverse transcription-PCR (qPCR) assays that detect influenza A/H3N2 virus RNA were reported and associated with two mutations (A37T and C161T) in the Matrix-encoding (M1) gene located on viral segment 7. This triggered a national alert in France. The present study sought to assess the association between the presence of these mutations and potential false negative results of influenza A/H3N2 virus RNA detection by commercialized qPCR assays at the clinical virology laboratory of our university hospitals in southern France. This study focused on the genetic diversity in the M1 gene and segment 7 of 624 influenza A/H3N2 virus genomes obtained from respiratory samples having tested qPCR-positive with M1 gene-targeting assays in our clinical virology laboratory. A total of 585 among the 624 influenza A/H3N2 virus genomes (93.7%) were of clade 3C.2a1b.2a.2, and 39 (6.3%) were of clade 3C.2a1b.1a. M1 gene substitutions A37T and C161T were both present in 582 (93.3%) genomes, only of clade 3C.2a1b.2a.2. Substitution A37T was present in 621 (99.5%) genomes. Substitution C161T was present in 585 genomes (93.8%), all of clade 3C.2a1b.2a.2. Moreover, 21 other nucleotide positions were mutated in ≥90% of the genomes. The present study shows that A37T/C and C161T mutations, and other mutations in the M1 gene and segment 7, were widely present in influenza A/H3N2 virus genomes recovered from respiratory samples diagnosed qPCR-positive with commercialized assays.

Influenza Incidence and Vaccine Effectiveness During the Southern Hemisphere Influenza Season - Chile, 2022.

The COVID-19 pandemic has affected influenza virus transmission, with historically low activity, atypical timing, or altered duration of influenza seasons during 2020-22 (1,2). Community mitigation measures implemented since 2020, including physical distancing and face mask use, have, in part, been credited for low influenza detections globally during the pandemic, compared with those during prepandemic seasons (1). Reduced population exposure to natural influenza infections during 2020-21 and relaxed community mitigation measures after introduction of COVID-19 vaccines could increase the possibility of severe influenza epidemics. Partners in Chile and the United States assessed Southern Hemisphere influenza activity and estimated age-group-specific rates of influenza-attributable hospitalizations and vaccine effectiveness (VE) in Chile in 2022. Chile's most recent influenza season began in January 2022, which was earlier than during prepandemic seasons and was associated predominantly with influenza A(H3N2) virus, clade 3C.2a1b.2a.2. The cumulative incidence of influenza-attributable pneumonia and influenza (P&I) hospitalizations was 5.1 per 100,000 person-years during 2022, which was higher than that during 2020-21 but lower than incidence during the 2017-19 influenza seasons. Adjusted VE against influenza A(H3N2)-associated hospitalization was 49%. These findings indicate that influenza activity continues to be disrupted after emergence of SARS-CoV-2 in 2020. Northern Hemisphere countries might benefit from preparing for an atypical influenza season, which could include early influenza activity with potentially severe disease during the 2022-23 season, especially in the absence of prevention measures, including vaccination. Health authorities should encourage all eligible persons to seek influenza vaccination and take precautions to reduce transmission of influenza (e.g., avoiding close contact with persons who are ill).

Influenza A viruses circulating in dogs: A review of the scientific literature.

Influenza A viruses (IAV) cause persistent epidemics and occasional human pandemics, leading to considerable economic losses. The ecology and epidemiology of IAV are very complex and the emergence of novel zoonotic pathogens is one of the greatest challenges in the healthcare. IAV are characterized by genetic and antigenic variability resulting from a combination of high mutation rates and a segmented genome that provides the ability to rapidly change and adapt to new hosts. In this context, available scientific evidence is of great importance for understanding the epidemiology and evolution of influenza viruses. The present review summarizes original research papers and IAV infections reported in dogs all over the world. Reports of interspecies transmission of equine influenza viruses H3N2 from birds to dogs, as well as double and triple reassortant strains resulting from reassortment of avian, human, and canine strains have amplified the genetic variety of canine influenza viruses. A total of 146 articles were deemed acceptable by PubMed and the Google Scholar database and were therefore included in this review. The largest number of research articles (n = 68) were published in Asia, followed by the Americas (n = 44), Europe (n = 31), Africa (n = 2), and Australia (n = 1). Publications are conventionally divided into three categories. The first category (largest group) included modern articles published from 2011 to the present (n = 93). The second group consisted of publications from 2000 to 2010 (n = 46). Single papers of 1919, 1931, 1963, 1972, 1975, and 1992 were also used, which was necessary to emphasize the history of the study of the ecology and evolution of the IAV circulating among various mammalian species. The largest number of publications occurred in 2010 (n = 18) and 2015 (n = 11), which is associated with IAV outbreaks observed at that time in the dog population in America, Europe, and Asia. In general, these findings raise concerns that dogs may mediate the adaptation of IAVs to zoonotic transmission and therefore serve as alternative hosts for genetic reassortment of these viruses. The global concern and significant threat to public health from the present coronavirus diseases 2019 pandemic confirms the necessity for active surveillance of zoonotic viral diseases with pandemic potential.

A general approach to identify low-frequency variants within influenza samples collected during routine surveillance.

Influenza viruses exhibit considerable diversity between hosts. Additionally, different quasispecies can be found within the same host. High-throughput sequencing technologies can be used to sequence a patient-derived virus population at sufficient depths to identify low-frequency variants (LFV) present in a quasispecies, but many challenges remain for reliable LFV detection because of experimental errors introduced during sample preparation and sequencing. High genomic copy numbers and extensive sequencing depths are required to differentiate false positive from real LFV, especially at low allelic frequencies (AFs). This study proposes a general approach for identifying LFV in patient-derived samples obtained during routine surveillance. Firstly, validated thresholds were determined for LFV detection, whilst balancing both the cost and feasibility of reliable LFV detection in clinical samples. Using a genetically well-defined population of influenza A viruses, thresholds of at least 104 genomes per microlitre and AF of ≥5 % were established as detection limits. Secondly, a subset of 59 retained influenza A (H3N2) samples from the 2016-2017 Belgian influenza season was composed. Thirdly, as a proof of concept for the added value of LFV for routine influenza monitoring, potential associations between patient data and whole genome sequencing data were investigated. A significant association was found between a high prevalence of LFV and disease severity. This study provides a general methodology for influenza LFV detection, which can also be adopted by other national influenza reference centres and for other viruses such as SARS-CoV-2. Additionally, this study suggests that the current relevance of LFV for routine influenza surveillance programmes might be undervalued.

Simultaneous Detection of RIG-1, MDA5, and IFIT-1 Expression Is a Convenient Tool for Evaluation of the Interferon-Mediated Response.

In this study, we developed a novel, multiplex qPCR assay for simultaneous detection of RIG-1, MDA5, and IFIT-1 at the mRNA level. The assay was validated in A549 cells transfected with in vitro transcribed RNAs. Both exogenous RNA-GFP and self-amplifying (saRNA-GFP) induced significant expression of RIG-1, MDA5, IFIT-1, as well as type I and III interferons. In contrast, native RNA from intact A549 cells did not upregulate expression of these genes. Next, we evaluated RIG-1, MDA5, and IFIT-1 mRNA levels in the white blood cells of patients with influenza A virus (H3N2) or SARS-CoV-2. In acute phase (about 4 days after disease onset) both viruses induced these genes expression. Clinical observations of SARS-CoV-2 typically describe a two-step disease progression, starting with a mild-to-moderate presentation followed by a secondary respiratory worsening 9 to 12 days after the first onset of symptoms. It revealed that the expression of RIG-1, MDA5, and MxA was not increased after 2 and 3 weeks from the onset the disease, while for IFIT-1 it was observed the second peak at 21 day post infection. It is well known that RIG-1, MDA5, and IFIT-1 expression is induced by the action of interferons. Due to the ability of SOCS-1 to inhibit interferon-dependent signaling, and the distinct antagonism of SARS-CoV-2 in relation to interferon-stimulated genes expression, we assessed SOCS-1 mRNA levels in white blood cells. SARS-CoV-2 patients had increased SOCS-1 expression, while the influenza-infected group did not differ from heathy donors. Moreover, SOCS-1 mRNA expression remained stably elevated during the course of the disease. It can be assumed that augmented SOCS-1 expression is one of multiple mechanisms that allow SARS-CoV-2 to escape from the interferon-mediated immune response. Our results implicate SOCS-1 involvement in the pathogenesis of SARS-CoV-2.

Development of an IoT-integrated multiplexed digital PCR system for quantitative detection of infectious diseases.

For rapid detection of the COVID-19 infection, the digital polymerase chain reaction (dPCR) with higher sensitivity and specificity has been presented as a promising method of point-of-care testing (POCT). Unlike the conventional real-time PCR (qPCR), the dPCR system allows absolute quantification of the target DNA without a calibration curve. Although a number of dPCR systems have previously been reported, most of these previous assays lack multiplexing capabilities. As different variants of COVID-19 have rapidly emerged, there is an urgent need for highly specific multiplexed detection systems. Additionally, the advances in the Internet of Things (IoT) technology have enabled the onsite detection of infectious diseases. Here, we present an IoT-integrated multiplexed dPCR (IM-dPCR) system involving sample compartmentalization, DNA amplification, fluorescence imaging, and quantitative analysis. This IM-dPCR system comprises three modules: a plasmonic heating-based thermal cycler, a multi-color fluorescence imaging set-up, and a firmware control module. Combined with a custom-developed smartphone application built on an IoT platform, the IM-dPCR system enabled automatic processing, data collection, and cloud storage. Using a self-priming microfluidic chip, 9 RNA groups (e.g., H1N1, H3N2, IFZ B, DENV2, DENV3, DENV4, OC43, 229E, and NL63) associated with three infectious diseases (e.g., influenza, dengue, and human coronaviruses) were analyzed with higher linearity (>98%) and sensitivity (1 copy per µL). The IM-dPCR system exhibited comparable analytical accuracy to commercial qPCR platforms. Therefore, this IM-dPCR system plays a crucial role in the onsite detection of infectious diseases.

Resurgence of Influenza Circulation in the Russian Federation during the Delta and Omicron COVID-19 Era.

Influenza circulation was substantially reduced after March 2020 in the European region and globally due to the wide introduction of non-pharmaceutical interventions (NPIs) against COVID-19. The virus, however, has been actively circulating in natural reservoirs. In summer 2021, NPIs were loosened in Russia, and influenza activity resumed shortly thereafter. Here, we summarize the epidemiological and virological data on the influenza epidemic in Russia in 2021-2022 obtained by the two National Influenza Centers. We demonstrate that the commonly used baseline for acute respiratory infection (ARI) is no longer sufficiently sensitive and BL for ILI incidence was more specific for early recognition of the epidemic. We also present the results of PCR detection of influenza, SARS-CoV-2 and other respiratory viruses as well as antigenic and genetic analysis of influenza viruses. Influenza A(H3N2) prevailed this season with influenza B being detected at low levels at the end of the epidemic. The majority of A(H3N2) viruses were antigenically and genetically homogenous and belonged to the clade 3C.2a1b.2a.2 of the vaccine strain A/Darwin/9/2021 for the season 2022-2023. All influenza B viruses belonged to the Victoria lineage and were similar to the influenza B/Austria/1359417/2021 virus. No influenza A(H1N1)pdm09 and influenza B/Yamagata lineage was isolated last season.

Evolution of the PB1 gene of human influenza A (H3N2) viruses circulating between 1968 and 2019.

One avian H3N2 influenza virus, providing its PB1 and HA segments, reassorted with one human H2N2 virus and caused a pandemic outbreak in 1968, killing over 1 million people. After its introduction to humanity, the pandemic H3N2 virus continued adapting to humans and has resulted in epidemic outbreaks every influenza season. To understand the functional roles of the originally avian PB1 gene in the circulating strains of human H3N2 influenza viruses, we analyzed the evolution of the PB1 gene in all human H3N2 isolates from 1968 to 2019. We found several specific residues dramatically changed around 2002-2009 and remained stable through to 2019. Then, we verified the functions of these PB1 mutations in the genetic background of the early pandemic virus, A/Hong Kong/1/1968(HK/68), as well as a recent seasonal strain, A/Jiangsu/34/2016 (JS/16). The PB1 V709I or PB1 V113A/K586R/D619N/V709I induced higher polymerase activity of HK/68 in human cells. And the four mutations acted cooperatively that had an increased replication capacity in vitro and in vivo at an early stage of infection. In contrast, the backward mutant, A113V/R586K/N619D/I709V, reduced polymerase activity in human cells. The PB1 I709V decreased viral replication in vitro, but this mutant only showed less effect on mice infection experiment, which suggested influenza A virus evolved in human host was not always consisted with highly replication efficiency and pathogenicity in other mammalian host. Overall, our results demonstrated that the identified PB1 mutations contributed to the viral evolution of human influenza A (H3N2) viruses.

Influenza Activity and Composition of the 2022-23 Influenza Vaccine - United States, 2021-22 Season.

Before the emergence of SARS-CoV-2, the virus that causes COVID-19, influenza activity in the United States typically began to increase in the fall and peaked in February. During the 2021-22 season, influenza activity began to increase in November and remained elevated until mid-June, featuring two distinct waves, with A(H3N2) viruses predominating for the entire season. This report summarizes influenza activity during October 3, 2021-June 11, 2022, in the United States and describes the composition of the Northern Hemisphere 2022-23 influenza vaccine. Although influenza activity is decreasing and circulation during summer is typically low, remaining vigilant for influenza infections, performing testing for seasonal influenza viruses, and monitoring for novel influenza A virus infections are important. An outbreak of highly pathogenic avian influenza A(H5N1) is ongoing; health care providers and persons with exposure to sick or infected birds should remain vigilant for onset of symptoms consistent with influenza. Receiving a seasonal influenza vaccine each year remains the best way to protect against seasonal influenza and its potentially severe consequences.

Rapid and universal detection of SARS-CoV-2 and influenza A virus using a reusable dual-channel optic fiber immunosensor.

Establishment of rapid on-site detection technology capable of concurrently detecting SARS-Cov-2 and influenza A virus is urgent to effectively control the epidemic from these two types of important viruses. Accordingly, we developed a reusable dual-channel optical fiber immunosensor (DOFIS), which utilized the evanescent wave-sensing properties and tandem detection mode of the mobile phase, effectively accelerating the detection process such that it can be completed within 10 min. It could detect the nucleoprotein of multiple influenza A viruses (H1N1, H3N2, and H7N9), as well as the spike proteins of the SARS-CoV-2 Omicron and Delta variants, and could respond to 20 TCID50 /ml SARS-CoV-2 pseudovirus and 100 TCID50 /ml influenza A (A/PR/8/H1N1), presenting lower limit of detection and wider linear range than enzyme-linked immunosorbent assay. The detection results on 26 clinical samples for SARS-CoV-2 demonstrated its specificity (100%) and sensitivity (94%), much higher than the sensitivity of commercial colloidal gold test strip (35%). Particularly, DOFIS might be reused more than 80 times, showing not only cost-saving but also potential in real-time monitoring of the pathogenic viruses. Therefore, this newly-developed DOFIS platform is low cost, simple to operate, and has broad spectrum detection capabilities for SARS-CoV-2 mutations and multiple influenza A strains. It may prove suitable for deployment as a rapid on-site screening and surveillance technique for infectious disease.

Sequence-Specific Features of Short Double-Strand, Blunt-End RNAs Have RIG-I- and Type 1 Interferon-Dependent or -Independent Anti-Viral Effects.

Pathogen-associated molecular patterns, including cytoplasmic DNA and double-strand (ds)RNA trigger the induction of interferon (IFN) and antiviral states protecting cells and organisms from pathogens. Here we discovered that the transfection of human airway cell lines or non-transformed fibroblasts with 24mer dsRNA mimicking the cellular micro-RNA (miR)29b-1* gives strong anti-viral effects against human adenovirus type 5 (AdV-C5), influenza A virus X31 (H3N2), and SARS-CoV-2. These anti-viral effects required blunt-end complementary RNA strands and were not elicited by corresponding single-strand RNAs. dsRNA miR-29b-1* but not randomized miR-29b-1* mimics induced IFN-stimulated gene expression, and downregulated cell adhesion and cell cycle genes, as indicated by transcriptomics and IFN-I responsive Mx1-promoter activity assays. The inhibition of AdV-C5 infection with miR-29b-1* mimic depended on the IFN-alpha receptor 2 (IFNAR2) and the RNA-helicase retinoic acid-inducible gene I (RIG-I) but not cytoplasmic RNA sensors MDA5 and ZNFX1 or MyD88/TRIF adaptors. The antiviral effects of miR29b-1* were independent of a central AUAU-motif inducing dsRNA bending, as mimics with disrupted AUAU-motif were anti-viral in normal but not RIG-I knock-out (KO) or IFNAR2-KO cells. The screening of a library of scrambled short dsRNA sequences identified also anti-viral mimics functioning independently of RIG-I and IFNAR2, thus exemplifying the diverse anti-viral mechanisms of short blunt-end dsRNAs.