Evidence of reassortment of avian influenza A (H2) viruses in Brazilian shorebirds.

Influenza A viruses of the H2 subtype represent a zoonotic and pandemic threat to humans due to a lack of widespread specific immunity. Although A(H2) viruses that circulate in wild bird reservoirs are distinct from the 1957 pandemic A(H2N2) viruses, there is concern that they could impact animal and public health. There is limited information on AIVs in Latin America, and next to nothing about H2 subtypes in Brazil. In the present study, we report the occurrence and genomic sequences of two influenza A viruses isolated from wild-caught white-rumped sandpipers (Calidris fuscicollis). One virus, identified as A(H2N1), was isolated from a bird captured in Restinga de Jurubatiba National Park (PNRJ, Rio de Janeiro), while the other, identified as A(H2N2), was isolated from a bird captured in Lagoa do Peixe National Park (PNLP, Rio Grande do Sul). DNA sequencing and phylogenetic analysis of the obtained sequences revealed that each virus belonged to distinct subtypes. Furthermore, the phylogenetic analysis indicated that the genomic sequence of the A(H2N1) virus isolated from PNRJ was most closely related to other A(H2N1) viruses isolated from North American birds. On the other hand, the A(H2N2) virus genome recovered from the PNLP-captured bird exhibited a more diverse origin, with some sequences closely related to viruses from Iceland and North America, and others showing similarity to virus sequences recovered from birds in South America. Viral genes of diverse origins were identified in one of the viruses, indicating local reassortment. This suggests that the extreme South of Brazil may serve as an environment conducive to reassortment between avian influenza virus lineages from North and South America, potentially contributing to an increase in overall viral diversity.

VP4 Mutation Boosts Replication of Recombinant Human/Simian Rotavirus in Cell Culture.

Rotavirus A (RVA) is the leading cause of diarrhea requiring hospitalization in children and causes over 100,000 annual deaths in Sub-Saharan Africa. In order to generate next-generation vaccines against African RVA genotypes, a reverse genetics system based on a simian rotavirus strain was utilized here to exchange the antigenic capsid proteins VP4, VP7 and VP6 with those of African human rotavirus field strains. One VP4/VP7/VP6 (genotypes G9-P[6]-I2) triple-reassortant was successfully rescued, but it replicated poorly in the first cell culture passages. However, the viral titer was enhanced upon further passaging. Whole genome sequencing of the passaged virus revealed a single point mutation (A797G), resulting in an amino acid exchange (E263G) in VP4. After introducing this mutation into the VP4-encoding plasmid, a VP4 mono-reassortant as well as the VP4/VP7/VP6 triple-reassortant replicated to high titers already in the first cell culture passage. However, the introduction of the same mutation into the VP4 of other human RVA strains did not improve the rescue of those reassortants, indicating strain specificity. The results show that specific point mutations in VP4 can substantially improve the rescue and replication of recombinant RVA reassortants in cell culture, which may be useful for the development of novel vaccine strains.

Phylogeography and gene pool analysis of highly pathogenic avian influenza H5N1 viruses reported in India from 2006 to 2021.

India has reported highly pathogenic avian influenza (HPAI) H5N1 virus outbreaks since 2006, with the first human case reported in 2021. These included viruses belonging to the clades 2.2, 2.2.2, 2.2.2.1, 2.3.2.1a, and 2.3.2.1c. There are currently no data on the gene pool of HPAI H5N1 viruses in India. Molecular clock and phylogeography analysis of the HA and NA genes; and phylogenetic analysis of the internal genes of H5N1 viruses from India were carried out. Sequences reported from 2006 to 2015; and sequences from 2021 that were available in online databases were used in the analysis. Five separate introductions of H5N1 viruses into India were observed, via Indonesia or Korea (2002), Bangladesh (2009), Bhutan (2010), and China (2013, 2018) (clades 2.2, 2.2.2, 2.2.2.1, 2.3.2.1a, 2.3.2.1c, and 2.3.4.4b). Phylogenetic analysis revealed eight reassortant genotypes. The H5N1 virus isolated from the human case showed a unique reassortant genotype. Amino acid markers associated with adaptation to mammals were also present. This is the first report of the spatio-temporal origins and gene pool analysis of H5N1 viruses from India, highlighting the need for increased molecular surveillance.

Evolution of H6N6 viruses in China between 2014 and 2019 involves multiple reassortment events.

H6N6 avian influenza viruses (AIVs) have been widely detected in wild birds, poultry, and even mammals. Recently, H6N6 viruses were reported to be involved in the generation of H5 and H7 subtype viruses. To investigate the emergence, evolutionary pattern, and potential for an epidemic of H6N6 viruses, the complete genomes of 198 H6N6 viruses were analyzed, including 168 H6N6 viruses deposited in the NCBI and GISAID databases from inception to January 2019 and 30 isolates collected from China between November 2014 and January 2019. Using phylogenetic analysis, the 198 strains of H6N6 viruses were identified as 98 genotypes. Molecular clock analysis indicated that the evolution of H6N6 viruses in China was constant and not interrupted by selective pressure. Notably, the laboratory isolates reassorted with six subtype viruses: H6N2, H5N6, H7N9, H5N2, H4N2, and H6N8, resulting in nine novel H6N6 reassortment events. These results suggested that H6N6 viruses can act as an intermediary in the evolution of H5N6, H6N6, and H7N9 viruses. Animal experiments demonstrated that the 10 representative H6N6 viruses showed low pathogenicity in chickens and were capable of infecting mice without prior adaptation. Our findings suggest that H6N6 viruses play an important role in the evolution of AIVs, and it is necessary to continuously monitor and evaluate the potential epidemic of the H6N6 subtype viruses.

[The identification of a novel reassortant H3N2 avian influenza virus based on nanopore sequencing technology and genetic characterization].

Objective: To identify a novel reassortant H3N2 avian influenza virus using nanopore sequencing technology and analyze its genetic characteristics. Methods: The positive samples of the H3N2 avian influenza virus, collected from the external environment in the farmers' market of Guangzhou, were cultured in chicken embryos. The whole genome was sequenced by targeted amplification and nanopore sequencing technology. The genetic characteristics were analyzed using bioinformatics software. Results: The phylogenetic trees showed that each gene fragment of the strain belonged to the Eurasian evolutionary branch, and the host source was of avian origin. The HA gene was closely related to the origin of the H3N6 virus. The NA gene was closely related to the H3N2 avian influenza virus from 2017 to 2020. The PB1 gene was closely related to the H5N6 avian influenza virus in Guangxi Zhuang Autonomous Region and Fujian Province from 2016 to 2022 and was not related to the PB1 gene of the H5N6 avian influenza epidemic strain in Guangzhou. The other internal gene fragments had complex sources with significant genetic diversity. Molecular characteristics indicated that the strain exhibited the molecular characteristics of a typical low pathogenic avian influenza virus and tended to bind to the receptors of avian origin. On important protein sites related to biological characteristics, this strain had mutations of PB2-L89V, PB1-L473V, NP-A184K, M1-N30D/T215A, and NS1-P42S/N205S. Conclusions: This study identified a novel reassortant H3N2 avian influenza virus by nanopore sequencing, with the PB1 gene derived from the H5N6 avian influenza virus. The virus had a low ability to spread across species, but further exploration was needed to determine whether its pathogenicity to the host was affected.

Long-term co-circulation of multiple influenza A viruses in pigs, Guangxi, China.

Influenza A viruses (IAVs) pose a persistent potential threat to human health because of the spillover from avian and swine infections. Extensive surveillance was performed in 12 cities of Guangxi, China, during 2018 and 2023. A total of 2540 samples (including 2353 nasal swabs and 187 lung tissues) were collected from 18 pig farms with outbreaks of respiratory disease. From these, 192 IAV-positive samples and 19 genomic sequences were obtained. We found that the H1 and H3 swine influenza A viruses (swIAVs) of multiple lineages and genotypes have continued to co-circulate during that time in this region. Genomic analysis revealed the Eurasian avian-like H1N1 swIAVs (G4) still remained predominant in pig populations. Strikingly, the novel multiple H3N2 genotypes were found to have been generated through the repeated introduction of the early H3N2 North American triple reassortant viruses (TR H3N2 lineage) that emerged in USA and Canada in 1998 and 2005, respectively. Notably, when the matrix gene segment derived from the H9N2 avian influenza virus was introduced into endemic swIAVs, this produced a novel quadruple reassortant H1N2 swIAV that could pose a potential risk for zoonotic infection.

Analyses and Insights into Genetic Reassortment and Natural Selection as Key Drivers of Piscine orthoreovirus Evolution.

Piscine orthoreovirus (PRV) is a pathogen that causes heart and skeletal muscle inflammation in Salmo salar and has also been linked to circulatory disorders in other farmed salmonids, such as Oncorhynchus kisutch and Oncorhynchus mykiss. The virus has a segmented, double-stranded RNA genome, which makes it possible to undergo genetic reassortment and increase its genomic diversity through point mutations. In this study, genetic reassortment in PRV was assessed using the full genome sequences available in public databases. This study used full genome sequences that were concatenated and genome-wide reassortment events, and phylogenetic analyses were performed using the recombination/reassortment detection program version 5 (RDP5 V 5.5) software. Additionally, each segment was aligned codon by codon, and overall mean distance and selection was tested using the Molecular Evolutionary Genetics Analysis X software, version 10.2 (MEGA X version 10.2). The results showed that there were 17 significant reassortment events in 12 reassortant sequences, involving genome exchange between low and highly virulent genotypes. PRV sequences from different salmonid host species did not appear to limit the reassortment. This study found that PRV frequently undergoes reassortment events to increase the diversity of its segmented genome, leading to antigenic variation and increased virulence. This study also noted that to date, no reassortment events have been described between PRV-1 and PRV-3 genotypes. However, the number of complete genomic sequences within each genotype is uneven. This is important because PRV-3 induces cross-protection against PRV-1, making it a potential vaccine candidate.

Abundant Intra-Subtype Reassortment Revealed in H13N8 Influenza Viruses.

Influenza A viruses (IAVs) pose a serious threat to global health. On the one hand, these viruses cause seasonal flu outbreaks in humans. On the other hand, they are a zoonotic infection that has the potential to cause a pandemic. The most important natural reservoir of IAVs are waterfowl. In this study, we investigated the occurrence of IAV in birds in the Republic of Buryatia (region in Russia). In 2020, a total of 3018 fecal samples were collected from wild migratory birds near Lake Baikal. Of these samples, 11 were found to be positive for the H13N8 subtype and whole-genome sequencing was performed on them. All samples contained the same virus with the designation A/Unknown/Buryatia/Arangatui-1/2020. To our knowledge, virus A/Unknown/Buryatia/Arangatui-1/2020 is the first representative of the H13N8 subtype collected on the territory of Russia, the sequence of which is available in the GenBank database. An analysis of reassortments based on the genome sequences of other known viruses has shown that A/Unknown/Buryatia/Arangatui-1/2020 arose as a result of reassortment. In addition, a reassortment most likely occurred several decades ago between the ancestors of the viruses recently collected in China, the Netherlands, the United States and Chile. The presence of such reassortment emphasizes the ongoing evolution of the H13N8 viruses distributed in Europe, North and East Asia, North and South America and Australia. This study underscores the importance of the continued surveillance and research of less-studied influenza subtypes.

Novel influenza A viruses in pigs with zoonotic potential, Chile.

Novel H1N2 and H3N2 swine influenza A viruses (IAVs) have recently been identified in Chile. The objective of this study was to evaluate their zoonotic potential. We perform phylogenetic analyses to determine the genetic origin and evolution of these viruses, and a serological analysis to determine the level of cross-protective antibodies in the human population. Eight genotypes were identified, all with pandemic H1N1 2009-like internal genes. H1N1 and H1N2 were the subtypes more commonly detected. Swine H1N2 and H3N2 IAVs had hemagglutinin and neuraminidase lineages genetically divergent from IAVs reported worldwide, including human vaccine strains. These genes originated from human seasonal viruses were introduced into the swine population since the mid-1980s. Serological data indicate that the general population is susceptible to the H3N2 virus and that elderly and young children also lack protective antibodies against the H1N2 strains, suggesting that these viruses could be potential zoonotic threats. Continuous IAV surveillance and monitoring of the swine and human populations is strongly recommended.IMPORTANCEIn the global context, where swine serve as crucial intermediate hosts for influenza A viruses (IAVs), this study addresses the pressing concern of the zoonotic potential of novel reassortant strains. Conducted on a large scale in Chile, it presents a comprehensive account of swine influenza A virus diversity, covering 93.8% of the country's industrialized swine farms. The findings reveal eight distinct swine IAV genotypes, all carrying a complete internal gene cassette of pandemic H1N1 2009 origin, emphasizing potential increased replication and transmission fitness. Genetic divergence of H1N2 and H3N2 IAVs from globally reported strains raises alarms, with evidence suggesting introductions from human seasonal viruses since the mid-1980s. A detailed serological analysis underscores the zoonotic threat, indicating susceptibility in the general population to swine H3N2 and a lack of protective antibodies in vulnerable demographics. These data highlight the importance of continuous surveillance, providing crucial insights for global health organizations.

Nucleoprotein reassortment enhanced transmissibility of H3 1990.4.a clade influenza A virus in swine.

The increased detection of H3 C-IVA (1990.4.a) clade influenza A viruses (IAVs) in US swine in 2019 was associated with a reassortment event to acquire an H1N1pdm09 lineage nucleoprotein (pdmNP) gene, replacing a TRIG lineage NP (trigNP). We hypothesized that acquiring the pdmNP conferred a selective advantage over prior circulating H3 viruses with a trigNP. To investigate the role of NP reassortment in transmission, we identified two contemporary 1990.4.a representative strains (NC/19 and MN/18) with different evolutionary origins of the NP gene. A reverse genetics system was used to generate wild-type (wt) strains and swap the pdm and TRIG lineage NP genes, generating four viruses: wtNC/19-pdmNP, NC/19-trigNP, wtMN/18-trigNP, and MN/18-pdmNP. The pathogenicity and transmission of the four viruses were compared in pigs. All four viruses infected 10 primary pigs and transmitted to five indirect contact pigs per group. Pigs infected via contact with MN/18-pdmNP shed virus 2 days earlier than pigs infected with wtMN/18-trigNP. The inverse did not occur for wtNC/19-pdmNP and NC/19-trigNP. This suggests that pdmNP reassortment resulted in a combination of genes that improved transmission efficiency when paired with the 1990.4.a hemagglutinin (HA). This is likely a multigenic trait, as replacing the trigNP gene did not diminish the transmission of a wild-type IAV in swine. This study demonstrates how reassortment and evolutionary change of internal genes can result in more transmissible viruses that influence HA clade detection frequency. Thus, rapidly identifying novel reassortants paired with dominant hemagglutinin/neuraminidase may improve the prediction of strains to include in vaccines.IMPORTANCEInfluenza A viruses (IAVs) are composed of eight non-continuous gene segments that can reassort during coinfection of a host, creating new combinations. Some gene combinations may convey a selective advantage and be paired together preferentially. A reassortment event was detected in swine in the United States that involved the exchange of two lineages of nucleoprotein (NP) genes (trigNP to pdmNP) that became a predominant genotype detected in surveillance. Using a transmission study, we demonstrated that exchanging the trigNP for a pdmNP caused the virus to shed from the nose at higher levels and transmit to other pigs more rapidly. Replacing a pdmNP with a trigNP did not hinder transmission, suggesting that transmission efficiency depends on interactions between multiple genes. This demonstrates how reassortment alters IAV transmission and that reassortment events can provide an explanation for why genetically related viruses with different internal gene combinations experience rapid fluxes in detection frequency.

The evolution, pathogenicity and transmissibility of quadruple reassortant H1N2 swine influenza virus in China: A potential threat to public health.

Swine are regarded as "intermediate hosts" or "mixing vessels" of influenza viruses, capable of generating strains with pandemic potential. From 2020 to 2021, we conducted surveillance on swine H1N2 influenza (swH1N2) viruses in swine farms located in Guangdong, Yunnan, and Guizhou provinces in southern China, as well as Henan and Shandong provinces in northern China. We systematically analyzed the evolution and pathogenicity of swH1N2 isolates, and characterized their replication and transmission abilities. The isolated viruses are quadruple reassortant H1N2 viruses containing genes from pdm/09 H1N1 (PB2, PB1, PA and NP genes), triple-reassortant swine (NS gene), Eurasian Avian-like (HA and M genes), and recent human H3N2 (NA gene) lineages. The NA, PB2, and NP of SW/188/20 and SW/198/20 show high gene similarities to A/Guangdong/Yue Fang277/2017 (H3N2). The HA gene of swH1N2 exhibits a high evolutionary rate. The five swH1N2 isolates replicate efficiently in human, canine, and swine cells, as well as in the turbinate, trachea, and lungs of mice. A/swine/Shandong/198/2020 strain efficiently replicates in the respiratory tract of pigs and effectively transmitted among them. Collectively, these current swH1N2 viruses possess zoonotic potential, highlighting the need for strengthened surveillance of swH1N2 viruses.

Isolation and characterization of genotype 4 Eurasian avian-like H1N1 influenza virus in pigs suffering from pneumonia.

Swine influenza viruses pose ongoing threat to pork industry throughout the world. In 2023, fattening pigs from a swine farm in Inner Mongolia of China experienced influenza-like symptoms. Co-infection of influenza A virus with Pasteurella multocida was diagnosed in lung tissues of diseased pigs and a genotype 4 (G4) Eurasian avian-like (EA) H1N1 virus was isolated, which was named as A/swine/Neimenggu/0326/2023. We demonstrated the virus preferentially bound human-like SAα2,6Gal receptor. It was noteworthy that the virus possessed multiple genetic markers for mammalian adaptation in the internal genes. Animal studies showed that compared with genotype 1 (G1) EA H1N1 virus and early prevalent G4 EA H1N1 virus, A/swine/Neimenggu/0326/2023 virus exhibited increased virus shedding, enhanced replication in lungs, and caused more severe lung lesions in pigs. These findings indicate that the G4 EA H1N1 virus poses increased threat to pork industry, controlling the prevailing viruses in pigs should be promptly implemented.

Evolution of influenza A viruses in exhibition swine and transmission to humans, 2013-2015.

AIMS: Swine are a mixing vessel for the emergence of novel reassortant influenza A viruses (IAV). Interspecies transmission of swine-origin IAV poses a public health and pandemic risk. In the United States, the majority of zoonotic IAV transmission events have occurred in association with swine exposure at agricultural fairs. Accordingly, this human-animal interface necessitates mitigation strategies informed by understanding of interspecies transmission mechanisms in exhibition swine. Likewise, the diversity of IAV in swine can be a source for novel reassortant or mutated viruses that pose a risk to both swine and human health. METHODS AND RESULTS: In an effort to better understand those risks, here we investigated the epidemiology of IAV in exhibition swine and subsequent transmission to humans by performing phylogenetic analyses using full genome sequences from 272 IAV isolates collected from exhibition swine and 23 A(H3N2)v viruses from human hosts during 2013-2015. Sixty-seven fairs (24.2%) had at least one pig test positive for IAV with an overall estimated prevalence of 8.9% (95% CI: 8.3-9.6, Clopper-Pearson). Of the 19 genotypes found in swine, 5 were also identified in humans. There was a positive correlation between the number of human cases of a genotype and its prevalence in exhibition swine. Additionally, we demonstrated that A(H3N2)v viruses clustered tightly with exhibition swine viruses that were prevalent in the same year. CONCLUSIONS: These data indicate that multiple genotypes of swine-lineage IAV have infected humans, and highly prevalent IAV genotypes in exhibition swine during a given year are also the strains detected most frequently in human cases of variant IAV. Continued surveillance and rapid characterization of IAVs in exhibition swine can facilitate timely phenotypic evaluation and matching of candidate vaccine strains to those viruses present at the human-animal interface which are most likely to spillover into humans.

Reassortant H9N2 canine influenza viruses containing the pandemic H1N1/2009 ribonucleoprotein complex circulating in pigs acquired enhanced virulence in mice.

The reassortment between avian H9N2 and Eurasian avian-like (EA) H1N1 viruses may have potentially changed from avian-to-mammals adaptation. This study generated 20 reassortant viruses with the introduction of H1N1/2009 internal genes from EA H1N1 virus into H9N2 virus. 12 of these recovered the replication capability both in the lungs and turbinate samples. 10 of 12 obtained PA gene segments from the ribonucleoprotein (RNP) complexes of the EA H1N1 virus, and 3 exhibited extreme virulence. Specially, the combination of PB2, PA and NP genes could overcome the species-specific restriction in human cells. Analysis of the polymerase activities found that introduction of the PA gene resulted in increased polymerase activity. These findings indicated that RNP complexes from EA H1N1 virus could confer an adaptation advantage and high compatibility to avian H9N2 virus. This raises new concerns for public health due to the possible coexistence of H9N2 and EA H1N1 viruses in dogs.

Hemagglutinin affects replication, stability and airborne transmission of the H9N2 subtype avian influenza virus.

H9N2 subtype avian influenza virus (AIV) can transmit by direct as well as airborne contacts. It has been widespread in poultry and continued to contribute to zoonotic spillover events by providing its six internal genes for the reassortment of novel influenza viruses (eg, H7N9) that infect poultry and humans. Compared to H7N9, H9N2 virus displays an efficient airborne transmissibility in poultry, but the mechanisms of transmission difference have been insufficiently studied. The Hemagglutinin (HA) and viral polymerase acidic protein (PA) have been implicated in the airborne transmission of influenza A viruses. Accordingly, we generated the reassortant viruses of circulating airborne transmissible H9N2 and non-airborne transmissible H7N9 viruses carrying HA and/or PA gene. The introduction of the PA gene from H7N9 into the genome of H9N2 virus resulted in a reduction in airborne transmission among chickens, while the isolated introduction of the HA gene segment completely eliminated airborne transmission among chickens. We further showed that introduction of HA gene of non-transmissible H7N9 did not influence the HA/NA balance of H9N2 virus, but increased the threshold for membrane fusion and decreased the acid stability. Thus, our results indicate that HA protein plays a key role in replication, stability, and airborne transmission of the H9N2 subtype AIV.

Experimental infection of pigs and ferrets with "pre-pandemic," human-adapted, and swine-adapted variants of the H1N1pdm09 influenza A virus reveals significant differences in viral dynamics and pathological manifestations.

Influenza A viruses are RNA viruses that cause epidemics in humans and are enzootic in the pig population globally. In 2009, pig-to-human transmission of a reassortant H1N1 virus (H1N1pdm09) caused the first influenza pandemic of the 21st century. This study investigated the infection dynamics, pathogenesis, and lesions in pigs and ferrets inoculated with natural isolates of swine-adapted, human-adapted, and "pre-pandemic" H1N1pdm09 viruses. Additionally, the direct-contact and aerosol transmission properties of the three H1N1pdm09 isolates were assessed in ferrets. In pigs, inoculated ferrets, and ferrets infected by direct contact with inoculated ferrets, the pre-pandemic H1N1pdm09 virus induced an intermediary viral load, caused the most severe lesions, and had the highest clinical impact. The swine-adapted H1N1pdm09 virus induced the highest viral load, caused intermediary lesions, and had the least clinical impact in pigs. The human-adapted H1N1pdm09 virus induced the highest viral load, caused the mildest lesions, and had the least clinical impact in ferrets infected by direct contact. The discrepancy between viral load and clinical impact presumably reflects the importance of viral host adaptation. Interestingly, the swine-adapted H1N1pdm09 virus was transmitted by aerosols to two-thirds of the ferrets. Further work is needed to assess the risk of human-to-human aerosol transmission of swine-adapted H1N1pdm09 viruses.

Emergence of Intergenogroup Reassortant G9P[4] Strains Following Rotavirus Vaccine Introduction in Ghana.

Rotavirus (RVA) is a leading cause of childhood gastroenteritis. RVA vaccines have reduced the global disease burden; however, the emergence of intergenogroup reassortant strains is a growing concern. During surveillance in Ghana, we observed the emergence of G9P[4] RVA strains in the fourth year after RVA vaccine introduction. To investigate whether Ghanaian G9P[4] strains also exhibited the DS-1-like backbone, as seen in reassortant G1/G3/G8/G9 strains found in other countries in recent years, this study determined the whole genome sequences of fifteen G9P[4] and two G2P[4] RVA strains detected during 2015-2016. The results reveal that the Ghanaian G9P[4] strains exhibited a double-reassortant genotype, with G9-VP7 and E6-NSP4 genes on a DS-1-like backbone (G9-P[4]-I2-R2-C2-M2-A2-N2-T2-E6-H2). Although they shared a common ancestor with G9P[4] DS-1-like strains from other countries, further intra-reassortment events were observed among the original G9P[4] and co-circulating strains in Ghana. In the post-vaccine era, there were significant changes in the distribution of RVA genotype constellations, with unique strains emerging, indicating an impact beyond natural cyclical fluctuations. However, reassortant strains may exhibit instability and have a limited duration of appearance. Current vaccines have shown efficacy against DS-1-like strains; however, ongoing surveillance in fully vaccinated children is crucial for addressing concerns about long-term effectiveness.