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The aim was to analyze heard immunity against influenza viruses as well as severe course of influenza infection prior to the 2019–2020 epidemic season. Methods. Blood sera samples were collected prior to and after conducting population-wide influenza vaccination campaign at the sanitary and epidemiological centers in different regions of the Russian Federation as well as at the Siberian Federal District, respectively. Sera samples were tested by using hemagglutination inhibition (HI) assay with vaccine strains A/Brisbane/02/2018 (H1N1)pdm09, A/Kansas/14/2017 (H3N2), B/Colorado/06/2017 (Victoria lineage). Baseline clinical and autopsy materials in case of influenza infection in vaccinated patients or severe and fatal influenza cases were collected to be tested by RT-PCR at the sanitary and epidemiological centers, Rospotrebnadzor. All influenza-virus positive samples were further sent to the SRC VB “Vector”. Results. A total of 7,896 and 600 blood serum samples were collected from subjects at Siberian Federal District prior to and after the populationwide influenza vaccination campaign, respectively. Prior to the epidemic season, the proportion of individuals seropositive for the influenza A virus subtypes A/(H1N1)pdm09 and A/H3N2 exceeded 50% in most of the regions, whereas frequency of those seropositive for the influenza B virus was profoundly lower ranging from 12 to 46% in the Northwestern Federal District and Volga Federal District, respectively. After influenza vaccination, the percentage of seropositive subjects in the Siberian Federal District increased as follows: for influenza subtype A/(H1N1)pdm09 — from 66 up to 79%, influenza subtype A/H3N2 — from 68 up to 78%, and for influenza B/Victoria — from 32 up to 47%. In 2019–2020, influenza B virus more frequently caused severe infection that agrees with the herd immunity data prior to the epidemic season. However, the vast majority of the influenza cases with fatal outcome was associated with influenza virus A A/H1N1pdm09 subtype. Conclusion. Quality of influenza vaccine, especially that one intended to vaccinate risk group subjects remains a crucial issue for contemporary scientific community. The study was conducted within the framework of the State Assignments no. 1/16 and 2/18.

—The receptor specificity for the A/rook/Chany/32/2015 (H5N1), A/common gull/Saratov/1676/2018 (H5N6), and A/chicken/PrimorskyKrai/03/2018 (Н9N2) strains of avian influenza virus has been studied. Using the method of surface plasmon resonance for the selected strains we obtained the values ​​of the equilibrium dissociation constants with analogues of receptors such as α2-3 and α2-6. It was found that for strains A/rook/Chany/32/2015 (H5N1) and A/common gull/Saratov/1676/2018 (H5N6), a binding with the human type of receptor (α2-6) is determined, in contrast to the strain A/chicken/PrimorskyKrai/03/2018 (H9N2), which is specific only for avian type receptors (α2-3).

Data obtained from monitoring cases of severe influenza, cases of vaccinated individuals, and unique cases were used to describe influenza viruses that circulated in Russia in the 2018–2019 epidemic season. A high proportion of the mutations D222G/N in A(H1N1)pdm09 HA was detected in fatal cases. Viruses of the B/Victoria lineage with deletions in HA were detected in Russia, and a reassortant seasonal influenza A(H1N2) virus was identified. A C-terminal truncation in the NS1 protein was detected in a substantial proportion of A(H3N2) viruses.

This study aimed to assess the antiviral susceptibility of influenza A(H5N8) viruses isolated in Russia in 2014–2018. Genetic analysis of 57 Russian isolates with full genome sequences did not find any markers of reduced susceptibility to baloxavir. Only one strain bore an amino acid substitution associated with adamantane resistance (M2-S31N). The neuraminidase of 1 strain had an NA-N293/294S (N8/N2 numbering) substitution associated with reduced inhibition by oseltamivir and normal inhibition by zanamivir, which was confirmed phenotypically. There were no other strains with reduced inhibition by oseltamivir and zanamivir in the phenotypic analysis. In order to estimate the worldwide prevalence of influenza A(H5N8) viruses bearing genetic markers of antiviral resistance, genome sequences deposited in the GISAID database were analyzed (database access: October 2020). The M2 protein of A(H5N8) viruses from the 2.3.4.4c clade had an M2-S31N substitution associated with reduced susceptibility to adamantanes. On the contrary, the majority (94%) of viruses from the 2.3.4.4b clade had the M2-S31 genotype. Fewer than 1% of analyzed viruses had amino acid substitutions associated with reduced susceptibility to baloxavir (PA-E199G, PA-E199E/G) or reduced or highly reduced inhibition by neuraminidase inhibitors (NA-R150/152K, NA-I221/222M, NA-I221/222I/M, NA-I221/222V, NA-I115/117V, NA-G145/147R, NA-R291/292R/K). An NA-N293/294S substitution was not present in sequences from the GISAID database. To the best of our knowledge, influenza A(H5N8) viruses with reduced inhibition by oseltamivir bearing an NA-N293/294S substitution have not been previously reported in epidemiological surveillance studies.

Aim. Control for the population herd immunity against seasonal influenza viruses as well as for emergence of antibodies against influenza with pandemic potential in human blood sera. Materials and methods. HAI reaction against vaccine and epidemic influenza viruses as well as HPAI viruses A/rook/Chany/32/2015 (H5N1) (clade 2.3.2.1c.) andA/Anhui/01/2013 (H7N9). Results. Among all the sera samples collected in the autumn of 2014 and 2015, none had reacted in HAI against A(H5N1) and A(H7N9) antigens even at 1:10 dilution. Among samples collected in autumn 2014, 41% were positive to A/Califorrna/07/09(HlNlpdm09) virus, 36% - A/Texas/50/2012 (H3N2), 40% - B/Brisbane/60/2008 (Vict.lin.) and 47% reacted in HAI against the B/Massachusetts/2/2012 (Yam.lin.) strain. 22% of all the samples had a titer of at least 40 against all the antigens and only 10% in HAI had a titer of 40 or more against all the vaccine strains. Among the samples collected in autumn 2015, the number of seropositive against A/Califorrna/07/09(HlNlpdm09) varied from 31% in the Urals FD to 46% in the Southern FD. The amount of seropositive against A/Switzerland/9715293/13 (H3N2) strain was at the level of 4 - 13% in all the FDs except Urals, where this parameter was slightly above 30%. The amount of seropositive against vaccine influenza В viruses varied from 23 to 76%. Only 2% of sera had titers in HAI of 40 or above against all the vaccine strains, 29% of all the samples were seronegative. Conclusion. Population immunity in Russia against influenza A(H3N2) is at a very low level, thus socially significant consequences of influenza epidemics in many aspects will depend on the vaccination campaign of autumn 2016.

In the spring of 2016, a loss of wild birds was observed during the monitoring of avian influenza virus activity in the Republic of Tyva. That outbreak was caused by influenza H5N8 virus of clade 2.3.4.4. In the fall, viruses of H5N8 clade 2.3.4.4 were propagated in European countries. This paper presents some results of analysis of the virus strains isolated during the spring and fall seasons in 2016 in the Russian Federation. The investigated strains were highly pathogenic for mice, and some of their antigenic and genetic features differed from those of an H5N8 strain that circulated in 2014 in Russia.

Neuraminidase (NA) thermostability of influenza A and B viruses isolated from birds, swine and humans was measured to evaluate its variability associated with host body temperature. The highest 50% inactivation temperature (IT50) was observed with H3N8 avian influenza virus (74 °C), and the lowest IT50 was observed with the seasonal human H3N2 virus (45.5 °C). The IT50 values of A(H1N1)pdm09 viruses 56.4-58.5 °C were statistically higher than that of the prepandemic strain A/Solomon Islands/03/06 (52.5 °C). An analysis of Ca2+ binding sites revealed the correspondence of amino acid changes to NA thermostability. This study demonstrates that changes in NA thermostability correspond to differences in host body temperature.

The 2017-2018 influenza epidemic season in Russia was characterized by a relatively low morbidity and mortality. We evaluated herd immunity prior to the 2017-2018 influenza season in hemagglutination inhibition assay, and performed characterization of influenza viruses isolated from severe or fatal influenza cases and from influenza cases in people vaccinated in the fall of 2017. During the 2017-2018 epidemic season, 87 influenza A and B viruses were isolated and viruses of the 75 influenza cases, including selected viral isolates and viruses analyzed directly from the original clinical material, were genetically characterized. The analyzed A(H1N1)pdm09 viruses belonged to clade 6B.1, B/Yamagata-like viruses belonged to clade 3, and B/Victoria-like viruses belonged to clade 1A and they were antigenically similar to the corresponding vaccine strains. A(H3N2) viruses belonged to clade 3C.2a and were difficult to characterize antigenically and the analysis indicated antigenic differences from the corresponding egg-grown vaccine strain. The next generation sequencing revealed the presence of D222/G/N polymorphism in the hemagglutinin gene in 32% of the analyzed A(H1N1)pdm09 lethal cases. This study demonstrated the importance of monitoring D222G/N polymorphism, including detection of minor viral variants with the mutations, in the hemagglutinin gene of A(H1N1)pdm09 for epidemiological surveillance. One strain of influenza virus A(H1N1)pdm09 was resistant to oseltamivir and had the H275Y amino acid substitution in the NA protein. All other isolates were susceptible to NA inhibitors. Prior to the 2017-2018 epidemic season, 67.4 million people were vaccinated, which accounted for 46.6% of the country's population. Just before the epidemic season 33-47% and 24-30% of blood sera samples collected within the territory of Russia showed the presence of protective antibody titers against vaccine strains of influenza A and influenza B/Victoria-like, respectively. Mass vaccination of the population had evidently reduced the severity of the flu epidemic during the 2017-2018 influenza epidemic season in Russia.

[This corrects the article DOI: 10.1371/journal.pone.0220401.].

Outbreaks of influenza, which is a contagious respiratory disease, occur throughout the world annually, affecting millions of people with many fatal cases. The D222G/N mutations in the hemagglutinin (HA) gene of A(H1N1)pdm09 are associated with severe and fatal human influenza cases. These mutations lead to increased virus replication in the lower respiratory tract (LRT) and may result in life-threatening pneumonia. Targeted NGS analysis revealed the presence of mutations in major and minor variants in 57% of fatal cases, with the proportion of viral variants with mutations varying from 1% to 98% in each individual sample in the epidemic season 2018–2019 in Russia. Co-occurrence of the mutations D222G and D222N was detected in a substantial number of the studied fatal cases (41%). The D222G/N mutations were detected at a low frequency (less than 1%) in the rest of the studied samples from fatal and nonfatal cases of influenza. The presence of HA D222Y/V/A mutations was detected in a few fatal cases. The high rate of occurrence of HA D222G/N mutations in A(H1N1)pdm09 viruses, their increased ability to replicate in the LRT and their association with fatal outcomes points to the importance of monitoring the mutations in circulating A(H1N1)pdm09 viruses for the evaluation of their epidemiological significance and for the consideration of disease prevention and treatment options.

This paper aims to characterize herd immunity of the population inhabiting Asian part of Russia before influenza epidemic and to describe influenza viruses isolated from severe cases including cases with fatal outcomes in the 2014-2015 epidemic season. HI test enabled us to study 3888 serum samples from healthy individuals including 1939 samples collected from poultry farm workers. We showed that none of the 3888 samples produced positive results with the antigens A/H5N1, А(H5N8) and A/H7N9. 41% of the samples are positive to A/California/07/09(H1N1pdm09), 36% of the samples are positive to A/Texas/50/2012 (H3N2), 40% of the samples are positive to B/Brisbane/60/2008 (Victoria lineage) and 47% of the samples are positive to B/Massachusetts/2/2012 (Yamagata lineage). In the 2014-2015 epidemic season 25 clinical and 19 autopsy samples were collected from individuals with severe flu-like infection. Fifteen influenza A(H3N2), two influenza A(H1N1pdm09) and one influenza B (Yamagata) virus strains were isolated in Madin-Darby Canine Kidney cell culture. All viruses exhibited normal inhibition by oseltamivir and zanamivir. A/KMAO/1/2015 and A/Kurgan/149/2015 were antigenically characterized as A/California/07/2009-like. Their hemagglutinin (HA) gene sequences fell into the predominant genetic group 6B and were similar to other recent H1N1pdm09 viruses circulating in Asian region. Eight H3N2 isolated viruses (A/Omsk/160/2015, A/Krasnoyarsk/324/2015, A/NizhnyNovgorod/788/2015, A/Omsk/141/2015, A/Buryatia/19/2015, A/Komi/99/2015, A/Novosibirsk/122/2015 and A/Chelyabinsk/160/2015) were characterized as A/Hong Kong/4801/2014-like and seven viruses (A/Irkutsk/88/2015, A/Krasnoyarsk/365/2015, A/Blagoveshchensk/19/2015, A/Kemerovo/2/2015, A/Chelyabinsk/160/2015, A/Chelyabinsk/192/2015 and A/Novosibirsk/64/2015) as A/Switzerland/9715293/2013-like. Their HA gene sequences belong to 3C.2a and 3C.3a genetic groups, respectively. B/Yekaterinburg/155/2015 virus was antigenically similar to B/Phuket/3073/2013 with HA sequence belonging to Y3 genetic group. Obtained findings are important for adjustment of public health measures and vaccine strategy in Russia.

In 2016-2017, several subtypes of the highly pathogenic avian influenza (HPAI) virus were isolated on the territory of Russia. In the autumn of 2016, during the avian influenza virus surveillance in the territory of the Kamchatka region of Russia the HPAI A(H5N5) influenza virus was isolated. Then, during 2016-2017, multiple outbreaks among wild birds and poultry caused by HPAI A(H5N8) avian influenza virus were recorded in European part of Russia. At the end of 2017, an outbreak among poultry caused by HPAI A(H5N2) influenza virus was recorded in the European part of Russia. Phylogenetic analysis of HA of the A(H5N5), A(H5N8), A(H5N2) showed the strains belong to the clade 2.3.4.4 b. All isolated strains were antigenically closely related to candidate vaccine viruses of clade 2.3.4.4 and showed high virulence in mice. Genetic analysis revealed presence of genetic markers potentially related to high virulence in mice in all studied viruses.

This work aimed to analyze the herd immunity to influenza among a Russian population living in regions with an increased risk of emergence of viruses with pandemic potential, and to isolate and investigate virus strains from severe influenza cases, including fatal cases, during the 2016-2017 epidemic season. In November 2016 - March 2017 highly pathogenic influenza outbreaks were registered in Russia among wild birds and poultry. No cases of human infection were registered. Analysis of 760 sera from people who had contact with infected or perished birds revealed the presence of antibodies to A(H5N1) virus of clade 2.3.2.1c and A(H5N8) virus of clade 2.3.4.4. The 2016-2017 influenza epidemic season in Russia began in weeks 46-47 of 2016 with predominant circulation of influenza A(H3N2) viruses. Strains isolated from severe influenza cases mainly belonged to 3C.2a.2 and 3C.2a.3 genetic groups. Up to the 8th week of 2017 severe influenza cases were often caused by influenza B viruses which belonged to 1A genetic group with antigenic properties similar to B/Brisbane/60/2008. All influenza A and B virus strains isolated in the 2016-2017 epidemic season were sensitive to oseltamivir and zanamivir.

The influenza epidemic in 2015–2016 in Russia is characterized by a sharp increase of influenza cases (beginning from the second week of 2016) with increased fatalities. Influenza was confirmed in 20 fatal cases registered among children (0–10 years), in 5 cases among pregnant women, and in 173 cases among elderly people (60 years and older). Two hundred and ninety nine people died from influenza were patients with some chronic problems. The overwhelming majority among the deceased (more than 98%) were not vaccinated against influenza. We isolated 109 influenza A(H1N1)pdm09 and one A(H3N2) virus strains from 501 autopsy material samples. The antigenic features of the strains were similar to the vaccine strains. A phylogenic analysis of hemagglutinin revealed that influenza A(H1N1)pdm09 virus strains belonged to 6B genetic group that had two main dominant subgroups during the 2015–2016 season. In Russia strains of the first group predominated. We registered an increased proportion of strains with D222G mutation in receptor-binding site. A herd immunity analysis carried out immediately prior to the epidemic showed that 34.4% blood sera samples collected in different regions of Russia were positive to A/California/07/09(H1N1)pdm09. We came to a conclusion that public awareness enhancement is necessary to reduce unreasonable refusals of vaccination.