N1 Gene Research Articles

Platelet gene expression and function in patients with COVID-19

AbstractThere is an urgent need to understand the pathogenesis of coronavirus disease 2019 (COVID-19). In particular, thrombotic complications in patients with COVID-19 are common and contribute to organ failure and mortality. Patients with severe COVID-19 present with hemostatic abnormalities that mimic disseminated intravascular coagulopathy associated with sepsis, with the major difference being increased risk of thrombosis rather than bleeding. However, whether severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection alters platelet function to contribute to the pathophysiology of COVID-19 remains unknown. In this study, we report altered platelet gene expression and functional responses in patients infected with SARS-CoV-2. RNA sequencing demonstrated distinct changes in the gene-expression profile of circulating platelets of COVID-19 patients. Pathway analysis revealed differential gene-expression changes in pathways associated with protein ubiquitination, antigen presentation, and mitochondrial dysfunction. The receptor for SARS-CoV-2 binding, angiotensin-converting enzyme 2 (ACE2), was not detected by messenger RNA (mRNA) or protein in platelets. Surprisingly, mRNA from the SARS-CoV-2 N1 gene was detected in platelets from 2 of 25 COVID-19 patients, suggesting that platelets may take-up SARS-COV-2 mRNA independent of ACE2. Resting platelets from COVID-19 patients had increased P-selectin expression basally and upon activation. Circulating platelet-neutrophil, -monocyte, and -T-cell aggregates were all significantly elevated in COVID-19 patients compared with healthy donors. Furthermore, platelets from COVID-19 patients aggregated faster and showed increased spreading on both fibrinogen and collagen. The increase in platelet activation and aggregation could partially be attributed to increased MAPK pathway activation and thromboxane generation. These findings demonstrate that SARS-CoV-2 infection is associated with platelet hyperreactivity, which may contribute to COVID-19 pathophysiology.

Validation of a single-step, single-tube reverse transcription loop-mediated isothermal amplification assay for rapid detection of SARS-CoV-2 RNA.

Introduction The SARS-CoV-2 pandemic of 2020 has resulted in unparalleled requirements for RNA extraction kits and enzymes required for virus detection, leading to global shortages. This has necessitated the exploration of alternative diagnostic options to alleviate supply chain issues.Aim To establish and validate a reverse transcription loop-mediated isothermal amplification (RT- LAMP) assay for the detection of SARS-CoV-2 from nasopharyngeal swabs.Methodology We used a commercial RT-LAMP mastermix from OptiGene in combination with a primer set designed to detect the CDC N1 region of the SARS-CoV-2 nucleocapsid (N) gene. A single-tube, single-step fluorescence assay was implemented whereby 1 µl of universal transport medium (UTM) directly from a nasopharyngeal swab could be used as template, bypassing the requirement for RNA purification. Amplification and detection could be conducted in any thermocycler capable of holding 65 °C for 30 min and measure fluorescence in the FAM channel at 1 min intervals.Results Assay evaluation by assessment of 157 clinical specimens previously screened by E-gene RT-qPCR revealed assay sensitivity and specificity of 87 and 100%, respectively. Results were fast, with an average time-to-positive (Tp) for 93 clinical samples of 14 min (sd±7 min). Using dilutions of SARS-CoV-2 virus spiked into UTM, we also evaluated assay performance against FDA guidelines for implementation of emergency-use diagnostics and established a limit-of-detection of 54 Tissue Culture Infectious Dose 50 per ml (TCID50 ml−1), with satisfactory assay sensitivity and specificity. A comparison of 20 clinical specimens between four laboratories showed excellent interlaboratory concordance; performing equally well on three different, commonly used thermocyclers, pointing to the robustness of the assay.Conclusion With a simplified workflow, The N1 gene Single Tube Optigene LAMP assay (N1-STOP-LAMP) is a powerful, scalable option for specific and rapid detection of SARS-CoV-2 and an additional resource in the diagnostic armamentarium against COVID-19.

Invitro diagnostics of coronavirus disease 2019: Technologies and application.

Laboratory-based diagnostic measures including virological and serological tests are essential for detecting severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Real-time reverse transcription-polymerase chain reactions (rRT-PCR) can detect SARS-COV-2 by targeting open reading frame-1 antibodies (ORF1ab), envelope protein, nucleocapsid protein, RNA-dependent RNA polymerase genes, and the N1, N2, and N3 (3N) target genes. Therefore, rRT-PCR remains the primary method of diagnosing SARS-CoV-2 despite being limited by false-negative results, long turnaround, complex protocols, and a need for skilled personnel. Serological diagnosis of coronavirus disease 2019 (COVID-19) is simple and does not require complex techniques and equipment, rendering it suitable for rapid detection and massive screening. However, serological tests cannot confirm SARS-CoV-2, and results will be false-negative when antibody concentrations fall below detection limits. Balancing the increased use of laboratory tests, risk of testing errors, need for tests, burden on healthcare systems, benefits of early diagnosis, and risk of unnecessary exposure is a significant and persistent challenge in diagnosing COVID-19.

Negligible effect of chicken cytokine IL-12 integration into recombinant fowlpox viruses expressing avian influenza virus neuraminidase N1 on host cellular immune responses.

In comparison to the extensive characterization of haemagglutinin antibodies of avian influenza virus (AIV), the role of neuraminidase (NA) as an immunogen is less well understood. This study describes the construction and cellular responses of recombinant fowlpox viruses (rFWPV) strain FP9, co-expressing NA N1 gene of AIV A/Chicken/Malaysia/5858/2004, and chicken IL-12 gene. Our data shows that the N1 and IL-12 proteins were successfully expressed from the recombinants with 48 kD and 70 kD molecular weights, respectively. Upon inoculation into specific-pathogen-free (SPF) chickens at 105 p.f.u. ml−1, levels of CD3+/CD4+ and CD3+/CD8+ populations were higher in the wild-type fowlpox virus FP9 strain, compared to those of rFWPV-N1 and rFWPV-N1-IL-12 at weeks 2 and 5 time points. Furthermore, rFWPV-N1-IL-12 showed a suppressive effect on chicken body weight within 4 weeks after inoculation. We suggest that co-expression of N1 with or without IL-12 offers undesirable quality as a potential AIV vaccine candidate.

A Novel DNA Vaccine Coding For H5 and N1 Genes of Highly Pathogenic Avian Influenza H5N1 Subtype

Control of avian influenza infection requires a good vaccine that could induce both humoral and cell-mediated immune response, specifically IFN-ɣ production, to maintain a high level of protection along with the minimal level of viral shedding after the infection to prevent secondary epidemics. In the current study, deoxyribonucleic Acid (DNA) vaccine coding for full-length H5 and N1 genes have been produced and evaluated in SPF-chicken. Humoral immune response estimated by haemagglutination inhibition (HI) assay revealed that the DNA vaccine gave a high titer of antibodies at the day 28-post vaccination and 14 days post-challenge. However, the shedding level was minimal with the DNA vaccine (0.1 Log 10 EID50). The IFN-ɣ transcript was upregulated at a higher level in the DNA vaccinated group. The results revealed that the DNA vaccine could induce a high level of humoral and IFN-ɣ level that maintains a high level of protection (92%) with the advantage of limiting the shedding level and thus, prevent secondary epidemics.

The Immunogenicity And Protective Efficacy Of Dna Vaccine Coding For Na1 Gene Of Highly Pathogenic Avian Influenza H5n1 Subtype

Control of avian influenza infection depends mainly on biosafety measures and vaccination, DNA vaccination is a novel method to generate antigen-specific antibody and cell-mediated immunity. In the current study, a DNA vaccine for a full-length N1 gene was developed in a trial to decrease the severity of the avian influenza virus spread and shedding. THE full-length N1 gene was cloned in entry clone p ENTER SD/TOPO, followed by homologous recombination with the destination mammalian expression vector (PDEST 40). The PDEST 40-N1 was used in the immunization of SPF chickens. Potency was evaluated through the survival rate that reaches 65%, which was far less than the commercially available inactivated vaccine. Meanwhile, the shedding of the virus from dead birds was 0.46 Log 10 EID50. At the same time, the most surprising result was the shedding level of the vaccinated live birds that were zero sheddings;on the other hand, the inactivated vaccine could not reduce the shedding level which remains very high (3.2 Log 10 EID50 ). IFN-γ transcript level in the DNA vaccinated group was detected by the 3rd-day post-vaccination and remained upregulated till the 28th post-vaccination. After the challenge, the level of IFN-γ was much higher until 14 days post-challenge. The inactivated vaccine could not stimulate any detectable level post-vaccination. These data suggested the ability of DNA vaccine coding for N1 gene of avian influenza to combat the virus shedding from live birds and could be used in combination with DNA vaccine coding for H5 to produce maximum protection with zero sheddings.

Virological surveillance of SARS-CoV-2 in an Italian northern area: comparison of Real Time RT PCR cycle threshold (Ct) values in three epidemic periods.

Aim of the study was to investigate the differences in Ct values in nasopharingeal swabs collected in three SARS-CoV-2 epidemic periods: first one from February 23 to March 25 (14 days from lockdown started on March 11); the second one from March 26 to May 18 (14 days from the end of strict lockdown on May 4) and the third one from May 19 until June 15. Viral RNA was detected in nasopharyngeal swabs obtained both from inpatients and outpatients. COVID-19 infection was confirmed according to the Ct values for N1 and N2 genes ascertained by Real-Time RT-PCR assay as described by the CDC. We calculated the prevalence of nasopharyngeal swabs tested positive for SARS-CoV-2, the mean and median of the Cts and the percentage of samples equal or below the Ct value of 25 in the 3 periods considered. The average value of Ct increased, going from 24.80 in the first epidemic period to 26.64 in the second period to 28.50 in the third period (p <0.001). The percentage of samples with Ct lower than or equal to 25 also decreased sharply from 54.7% to 20.0%. These findings need to be integrated with epidemiological and clinical data. (www.actabiomedica.it)

Evaluating the use of posterior oropharyngeal saliva in a point-of-care assay for the detection of SARS-CoV-2

ABSTRACT During the Coronavirus disease 2019 (COVID-19) pandemic, logistic problems associated with specimen collection limited the SARS-CoV-2 testing, especially in the community. In this study, we assessed the use of posterior oropharyngeal saliva as specimens for the detection of SARS-CoV-2 in an automated point-of-care molecular assay. Archived nasopharyngeal swab (NPS) and posterior oropharyngeal saliva specimens of 58 COVID-19 patients were tested with the Xpert® Xpress SARS-CoV-2 assay. SARS-CoV-2 was detected in either NPS or saliva specimens of all patients. Among them, 84.5% (49/58) tested positive in both NPS and saliva, 10.3% (6/58) tested positive in NPS only, and 5.2% (3/58) tested positive in saliva only. No significant difference in the detection rate was observed between NPS and saliva (McNemar’s test p = 0.5078). The detection rate was slightly higher for N2 (NPS 94.8% and Saliva 93.1%) than that of the E gene target (Saliva: 89.7% vs 82.8%) on both specimen types. Significantly earlier median Ct value was observed for NPS comparing to that of saliva on both E (26.8 vs 29.7, p = 0.0002) and N2 gene target (29.3 vs 32.3, p = 0.0002). The median Ct value of E gene target was significantly earlier than that of the N2 gene target for both NPS (26.8 vs 29.3, p < 0.0001) and saliva (29.7 vs 32.3, p < 0.0001). In conclusion, posterior oropharyngeal saliva and NPS were found to have similar detection rates in the point-of-care test for SARS-CoV-2 detection. Since posterior oropharyngeal saliva can be collected easily, the use of saliva as an alternative specimen type for SARS-CoV-2 detection is recommended.

Recompleting the Caenorhabditis elegans genome

Caenorhabditis elegans was the first multicellular eukaryotic genome sequenced to apparent completion. Although this assembly employed a standard C. elegans strain (N2), it used sequence data from several laboratories, with DNA propagated in bacteria and yeast. Thus, the N2 assembly has many differences from any C. elegans available today. To provide a more accurate C. elegans genome, we performed long-read assembly of VC2010, a modern strain derived from N2. Our VC2010 assembly has 99.98% identity to N2 but with an additional 1.8 Mb including tandem repeat expansions and genome duplications. For 116 structural discrepancies between N2 and VC2010, 97 structures matching VC2010 (84%) were also found in two outgroup strains, implying deficiencies in N2. Over 98% of N2 genes encoded unchanged products in VC2010; moreover, we predicted ≥53 new genes in VC2010. The recompleted genome of C. elegans should be a valuable resource for genetics, genomics, and systems biology.

Phenotypic and genotypic detection of microcin produced by some Enterobacteraceae

Back ground: Microcins are ribosomal synthesized antimicrobial peptides produced principally by bacteria of the Enterobacteriaceae family that are active against other bacteria, either in the same species (narrow spectrum), or across genera (broad spectrum) that share the same ecological niche.&#x0D; Objective: The ability of the Enterobacteriaceae to produce microsines.&#x0D; Materials and methods: One hundred urine samples have been collected from patients suffering from UTI whom admitted to Al Hakim Hospital in AL-Najaf City during the period from 17October 2017 to 13 February 2018.&#x0D; Results: The results of PCR technique for amplification show that all isolates were possess mic N by appearance of amplicon with molecular weight 938 bp while 46 (76%) isolates were possess mic N1and 24 (40%) isolates were possess mic N2 gene by appearance of amplicon with molecular weight 368 bp and 167 bp respectively.Cross-streak activity assay was used to detect the selected strains with antibacterial activities against pathogenic bacteria.&#x0D; Conclusion: some isolates have one or more gene microcin encoded of , however, microcin N is present in all isolates.

Differential gene expression of Thy0517 cells in thymoma after down-regulation of Wnt4 and fork head gene family protein N1 gene

Differential gene expression of Thy0517 cells in thymoma after down-regulation of Wnt4 and fork head gene family protein N1 gene

Appearance of reassortant European avian-origin H1 influenza A viruses of swine in Vietnam.

Three subtypes-H1N1, H1N2 and H3N2-of influenza A viruses of swine (IAVs-S) are currently endemic in swine worldwide, but there is considerable genotypic diversity among each subtype and limited geographical distribution. Through IAVs-S monitoring in Vietnam, two H1N2 influenza A viruses were isolated from healthy pigs in Ba Ria-Vung Tau Province, Southern Vietnam, on 2 December 2016. BLAST and phylogenetic analyses revealed that their HA and NA genes were derived from those of European avian-like H1N2 IAVs-S that contained avian-origin H1 and human-like N2 genes, and were particularly closely related to those of IAVs-S circulating in the Netherlands, Germany or Denmark. In addition, the internal genes of these Vietnamese isolates were derived from human A(H1N1)pdm09 viruses, suggesting that the Vietnamese H1N2 IAVs-S are reassortants between European H1N2 IAVs-S and human A(H1N1)pdm09v. The appearance of European avian-like H1N2 IAVs-S in Vietnam marks their first transmission outside Europe. Our results and statistical analyses of the number of live pigs imported into Vietnam suggest that the European avian-like H1N2 IAVs-S may have been introduced into Vietnam with their hosts through international trade. These findings highlight the importance of quarantining imported pigs to impede the introduction of new IAVs-S.

Multiplex real-time reverse transcription polymerase chain reaction for differential detection of H5, N1, and N8 genes of highly pathogenic avian influenza viruses

Rapid and differential diagnosis of highly pathogenic avian influenza virus (HPAIV) subtype H5 is essential for the effective prevention and control of outbreaks caused by this pathogen. In this study, we describe a one-step multiplex real-time reverse transcription polymerase chain reaction (mRRT-PCR), using H5-, N1-, and N8-specific primers and probes, for differential detection of two HPAIVs (H5N1 and H5N8) and other H5-subtype AIVs. Using the mRRT-PCR assay, we were able to detect H5N1, H5N8, and other H5-subtype AIVs in a one-tube reaction, with high specificity; furthermore, using an in silico PCR program, we confirmed that this assay can detect nearly all H5, N1, and N8 genes of AIVs currently available in the Influenza Sequence Database. The limit of detection of the assay was determined to be as low as 100 copies/reaction for each target gene, and was comparable to limits of detection of previously reported mRRT-PCR assays. Thus, the mRRT-PCR assay described here can serve as a rapid and reliable differential diagnostic tool for the monitoring and surveillance of H5N1, H5N8, and other H5-subtype AIVs in countries where these pathogens are problematic.

Molecular characterization of neuraminidase genes of influenza A(H3N2) viruses circulating in Southwest India from 2009 to 2013.

Molecular characterization of neuraminidase (NA) gene of 25 influenza A(H3N2) virus isolates (2009-2013) archived at the Manipal Centre for Virus Research was carried out. The annual rate of amino acid substitutions in the N2 gene of influenza A(H3N2) virus isolates was 0.2-0.6%. Out of the 25 NA sequences analyzed, catalytic site mutations were observed in three isolates. Two of the mutations (D151G and E276G) were detected in functional catalytic residues, and an E227V mutation was detected in the framework residues. To the best of our knowledge, NA inhibitor resistance associated with the mutations E276G and E227V has not been reported. However, the mutation D151G, which is commonly associated with culturing of influenza A(H3N2) virus in Madin-Darby canine kidney (MDCK) cells, has been reported to result in a reduction in virus susceptibility to NA inhibitor drugs. Our study also detected mutations in antigenic residues. Some of the mutations (except D197G, K249E, A250T, S334C, and H347R/N) remained conserved in isolates of succeeding seasons. Antigenic residue mutations (D197G and S334C) have not been reported globally to date. The effect of these catalytic and antigenic mutant residues on drug and antibody binding was analyzed using three-dimensional structural analysis and biochemical assays. Antigenic variability of influenza A(H3N2) viruses is a major concern, and vaccine failures are mainly due to genetic variations in the HA gene. Our study documents that genetic changes in N2 occur at a slower rate, and this information is useful for the consideration and standardization of NA in influenza vaccines.

The Establishment of Realtime Fluorescent Quantitative Polymerase chain reaction (PCR) for Detection of Highly Pathogenic Avian Influenza Virus Subtype H5N1

Highly pathogenic strains of avian influenza virus (AIV), which are influenza A viruses, cause severe disease in domestic poultry and humans. The objective of this study was to establish a fluorescent quantitative RT-PCR assay for detection of highly pathogenic avian influenza virus (AIV) subtype H5N1. The H5 and N1 subtypespecific probe sets were developed based on avian influenza virus sequences detected in China. Two pairs of primers and two fluorescent probes were strictly designed and optimized in a reaction system. According to the amount of plasmid RNA extracted from H5N1 strains, the standard curve DWQBGWDWQBGW of fluorescent quantitative PCR was drawn and all of the specimens were then tested by means of Real-time PCR. The test of highly pathogenic AIV subtype H5N1 was identified to be specific and its sensitivity level was 102~103 copies/reaction. The standard curve was accomplished at 109～105 DNA copies/reaction. It took only three hours from viral RNA extraction through to completion of the test. The assay was easy to carry out and highly reproducible. In conclusion, fluorescent quantitative PCR, described here, provides a rapid, specific and sensitive method to detect not only the H5 but N1 genes as well.

Genetic characterization of H5N2 influenza viruses isolated from wild birds in Japan suggests multiple reassortment.

Low-pathogenic avian influenza viruses (LPAIVs) of the H5 subtype can mutate to highly pathogenic forms, potentially destabilizing the poultry industry. Wild migratory birds are considered a natural reservoir of LPAIVs capable of dispersing both high- and low-pathogenic forms of the virus. Therefore, surveillance and characterization of AIV in wild birds are essential. Here, we report on the isolation and genetic characterization of 10 AIVs of the H5N2 subtype obtained through surveillance in Hokkaido, Japan, during 2009 and 2011. Full-genome sequencing revealed that the H5 and N2 genes of these isolates are all closely related to each other, belonging to the Eurasian avian-like lineage, but they are unrelated to H5 highly pathogenic strains of clade 2.3.4.4. The internal genes of the isolates were found to be diverse, consistent with our hypothesis that these H5N2 strains have undergone multiple reassortment events. Even though all of the H5N2 isolates were characterized as LPAIV based on the amino acid sequences at the HA cleavage site, this analysis demonstrates a diverse pool of precursors that may seed future outbreaks in poultry and possible human transmissions, suggesting the need for high-quality surveillance.

An immunoassay-based reverse-transcription loop-mediated isothermal amplification assay for the rapid detection of avian influenza H5N1 virus viremia

Avian influenza virus (AIV) subtype H5N1 attracts particular consideration because it is a continuous threat to animals and public health systems. The viremia caused by AIV H5N1 infection may increase the risk of blood-borne transmission between humans. Therefore, there is a need to rapidly evaluate and implement screening measures for AIV H5N1 viremia that allows for rapid response to this potentially pandemic threat. The present report describes an immunoassay-based reverse-transcription loop-mediated isothermal amplification (immuno-RT-LAMP) assay for the rapid detection of AIV H5N1 in whole blood samples. Using PCR tubes coated with an H5 subtype monoclonal antibody, AIV H5N1 virions were specifically captured from blood samples. After a thermal lysis step, the released viral N1 gene was exponentially amplified using RT-LAMP on either a real-time PCR instrument for quantitative analysis, or in a water bath system for endpoint analysis. The detection limit of the newly developed immuno-RT-LAMP assay was as low as 1.62×101 50% embryo infectious dose/mL of virus in both regular samples and simulated viremia samples. There were no cross-reactions with non-H5N1 influenza viruses or other avian viruses. The reproducibility of the assay was confirmed using intra- and inter-assay tests with variability ranging from 1.05% to 3.37%. Our results indicate that immuno-RT-LAMP is a novel, effective point-of-care virus identification solution for the rapid diagnosis and monitoring of AIV H5N1 in blood samples.

Novel Avian Influenza A(H5N8) Viruses in Migratory Birds, China, 2013-2014.

Novel Avian Influenza A(H5N8) Viruses in Migratory Birds, China, 2013-2014.

Widespread of H5N1 infections in apparently healthy backyard poultry.

Highly pathogenic avian influenza subtype H5N1 represents a threat to the poultry industry and human health worldwide. Inapparently infected birds are suspected to play an essential role in the spread of avian influenza virus. In the current study, a total of 25,646 samples (16,185 chicken, 4696 ducks, 1633 geese and 3132 turkeys) from apparently healthy birds were screened for the presence of positive samples for H5N1 during 2009-2014. The samples were examined by reverse transcriptase real-time polymerase chain reaction (rRT-PCR) for M, H5 and N1 genes of avian influenza viruses. The results revealed that the HPAI H5N1 existed in an inapparent manner in ducks (4.68%), geese (4.10%), chickens (2.48%) and turkeys (2.29%). The current finding highlights the serious impact of such type on birds in the epidemiology of H5N1 in birds, animals and humans. It also highlights the existence of another reason other than vaccination that contributes to the widespread of inapparent infection of H5N1 in Egypt.

Pathogenicity and transmissibility of novel reassortant H3N2 influenza viruses with 2009 pandemic H1N1 genes in pigs.

At least 10 different genotypes of novel reassortant H3N2 influenza viruses with 2009 pandemic H1N1 [A(H1N1)pdm09] gene(s) have been identified in U.S. pigs, including the H3N2 variant with a single A(H1N1)pdm09 M gene, which has infected more than 300 people. To date, only three genotypes of these viruses have been evaluated in animal models, and the pathogenicity and transmissibility of the other seven genotype viruses remain unknown. Here, we show that three H3N2 reassortant viruses that contain 3 (NP, M, and NS) or 5 (PA, PB2, NP, M, and NS) genes from A(H1N1)pdm09 were pathogenic in pigs, similar to the endemic H3N2 swine virus. However, the reassortant H3N2 virus with 3 A(H1N1)pdm09 genes and a recent human influenza virus N2 gene was transmitted most efficiently among pigs, whereas the reassortant H3N2 virus with 5 A(H1N1)pdm09 genes was transmitted less efficiently than the endemic H3N2 virus. Interestingly, the polymerase complex of reassortant H3N2 virus with 5 A(H1N1)pdm09 genes showed significantly higher polymerase activity than those of endemic and reassortant H3N2 viruses with 3 A(H1N1)pdm09 genes. Further studies showed that an avian-like glycine at position 228 at the hemagglutinin (HA) receptor binding site is responsible for inefficient transmission of the reassortant H3N2 virus with 5 A(H1N1)pdm09 genes. Taken together, our results provide insights into the pathogenicity and transmissibility of novel reassortant H3N2 viruses in pigs and suggest that a mammalian-like serine at position 228 in the HA is critical for the transmissibility of these reassortant H3N2 viruses. Swine influenza is a highly contagious zoonotic disease that threatens animal and public health. Introduction of 2009 pandemic H1N1 virus [A(H1N1)pdm09] into swine herds has resulted in novel reassortant influenza viruses in swine, including H3N2 and H1N2 variants that have caused human infections in the United States. We showed that reassortant H3N2 influenza viruses with 3 or 5 genes from A(H1N1)pdm09 isolated from diseased pigs are pathogenic and transmissible in pigs, but the reassortant H3N2 virus with 5 A(H1N1)pdm09 genes displayed less efficient transmissibility than the endemic and reassortant H3N2 viruses with 3 A(H1N1)pdm09 genes. Further studies revealed that an avian-like glycine at the HA 228 receptor binding site of the reassortant H3N2 virus with 5 A(H1N1)pdm09 genes is responsible for less efficient transmissibility in pigs. Our results provide insights into viral pathogenesis and the transmission of novel reassortant H3N2 viruses that are circulating in U.S. swine herds and warrant future surveillance.