Impact of Reinfection with SARS-CoV-2 Omicron Variants in Previously Infected Hamsters.

The diversity of SARS-CoV-2 mutations raises the possibility of reinfection of individuals previously infected with earlier variants, and this risk is further increased by the emergence of the B.1.1.529 Omicron variant. In this study, we used an in vivo, hamster infection model to assess the potential for individuals previously infected with SARS-CoV-2 to be reinfected with Omicron variant and we also investigated the pathology associated with such infections. Initially, Syrian hamsters were inoculated with a lineage A, B.1.1.7, B.1.351, B.1.617.2 or a subvariant of Omicron, BA.1 strain and then reinfected with the BA.1 strain 5 weeks later. Subsequently, the impact of reinfection with Omicron subvariants (BA.1 and BA.2) in individuals previously infected with the BA.1 strain was examined. Although viral infection and replication were suppressed in both the upper and lower airways, following reinfection, virus-associated RNA was detected in the airways of most hamsters. Viral replication was more strongly suppressed in the lower respiratory tract than in the upper respiratory tract. Consistent amino acid substitutions were observed in the upper respiratory tract of infected hamsters after primary infection with variant BA.1, whereas diverse mutations appeared in hamsters reinfected with the same variant. Histopathology showed no acute pneumonia or disease enhancement in any of the reinfection groups and, in addition, the expression of inflammatory cytokines and chemokines in the airways of reinfected animals was only mildly elevated. These findings are important for understanding the risk of reinfection with new variants of SARS-CoV-2. IMPORTANCE The emergence of SARS-CoV-2 variants and the widespread use of COVID-19 vaccines has resulted in individual differences in immune status against SARS-CoV-2. A decay in immunity over time and the emergence of variants that partially evade the immune response can also lead to reinfection. In this study, we demonstrated that, in hamsters, immunity acquired following primary infection with previous SARS-CoV-2 variants was effective in preventing the onset of pneumonia after reinfection with the Omicron variant. However, viral infection and multiplication in the upper respiratory tract were still observed after reinfection. We also showed that more diverse nonsynonymous mutations appeared in the upper respiratory tract of reinfected hamsters that had acquired immunity from primary infection. This hamster model reveals the within-host evolution of SARS-CoV-2 and its pathology after reinfection, and provides important information for countermeasures against diversifying SARS-CoV-2 variants.

A community cluster of influenza A(H3N2) virus infection with reduced susceptibility to baloxavir due to a PA E199G substitution in Japan, February to March 2023.

A community cluster of influenza A(H3N2) caused by viruses with an E199G substitution in PA was detected in Nara, Japan, between February and March 2023. The three patients with these mutant viruses had not received antiviral treatment before specimen collection but patients in the same hospital had. The sequences of the mutant viruses were closely related, suggesting clonal spread in Nara. They showed reduced susceptibility to baloxavir in vitro; however, the clinical significance of the PA E199G substitution remains unclear.

Human-to-Human Transmission of Influenza A(H3N2) Virus with Reduced Susceptibility to Baloxavir, Japan, February 2019.

In 2019, influenza A(H3N2) viruses carrying an I38T substitution in the polymerase acidic gene, which confers reduced susceptibility to baloxavir, were detected in Japan in an infant without baloxavir exposure and a baloxavir-treated sibling. These viruses' whole-genome sequences were identical, indicating human-to-human transmission. Influenza virus isolates should be monitored for baloxavir susceptibility.

Pathogenicity of two novel human-origin H7N9 highly pathogenic avian influenza viruses in chickens and ducks.

Human infection by low-pathogenic avian influenza viruses of the H7N9 subtype was first reported in March 2013 in China. Subsequently, these viruses caused five outbreaks through September 2017. In the fifth outbreak, H7N9 virus possessing a multiple basic amino acid insertion in the cleavage site of hemagglutinin emerged and caused 4% of all human infections in that period. To date, H7N9 highly pathogenic avian influenza viruses (HPAIVs) have been isolated from poultry, mostly chickens, as well as the environment. To evaluate the relative infectivity of these viruses in poultry, chickens and ducks were subjected to experimental infection with two H7N9 HPAIVs isolated from humans, namely A/Guangdong/17SF003/2016 and A/Taiwan/1/2017. When chickens were inoculated with the HPAIVs at a dose of 106 50% egg infectious dose (EID50), all chickens died within 2-5 days after inoculation, and the viruses replicated in most of the internal organs examined. The 50% lethal doses of A/Guangdong/17SF003/2016 and A/Taiwan/1/2017 in chickens were calculated as 103.3 and 104.7 EID50, respectively. Conversely, none of the ducks inoculated with either virus displayed any clinical signs, and less-efficient virus replication and less shedding were observed in ducks compared to chickens. These findings indicate that chickens, but not ducks, are highly permissive hosts for emerging H7N9 HPAIVs.

Characterization of H7N9 influenza A viruses isolated from humans.

Avian influenza A viruses rarely infect humans; however, when human infection and subsequent human-to-human transmission occurs, worldwide outbreaks (pandemics) can result. The recent sporadic infections of humans in China with a previously unrecognized avian influenza A virus of the H7N9 subtype (A(H7N9)) have caused concern owing to the appreciable case fatality rate associated with these infections (more than 25%), potential instances of human-to-human transmission, and the lack of pre-existing immunity among humans to viruses of this subtype. Here we characterize two early human A(H7N9) isolates, A/Anhui/1/2013 (H7N9) and A/Shanghai/1/2013 (H7N9); hereafter referred to as Anhui/1 and Shanghai/1, respectively. In mice, Anhui/1 and Shanghai/1 were more pathogenic than a control avian H7N9 virus (A/duck/Gunma/466/2011 (H7N9); Dk/GM466) and a representative pandemic 2009 H1N1 virus (A/California/4/2009 (H1N1pdm09); CA04). Anhui/1, Shanghai/1 and Dk/GM466 replicated well in the nasal turbinates of ferrets. In nonhuman primates, Anhui/1 and Dk/GM466 replicated efficiently in the upper and lower respiratory tracts, whereas the replicative ability of conventional human influenza viruses is typically restricted to the upper respiratory tract of infected primates. By contrast, Anhui/1 did not replicate well in miniature pigs after intranasal inoculation. Critically, Anhui/1 transmitted through respiratory droplets in one of three pairs of ferrets. Glycan arrays showed that Anhui/1, Shanghai/1 and A/Hangzhou/1/2013 (H7N9) (a third human A(H7N9) virus tested in this assay) bind to human virus-type receptors, a property that may be critical for virus transmissibility in ferrets. Anhui/1 was found to be less sensitive in mice to neuraminidase inhibitors than a pandemic H1N1 2009 virus, although both viruses were equally susceptible to an experimental antiviral polymerase inhibitor. The robust replicative ability in mice, ferrets and nonhuman primates and the limited transmissibility in ferrets of Anhui/1 suggest that A(H7N9) viruses have pandemic potential.

Detection of influenza A(H3N2) viruses exhibiting reduced susceptibility to the novel cap-dependent endonuclease inhibitor baloxavir in Japan, December 2018.

The novel cap-dependent endonuclease inhibitor baloxavir marboxil was approved for the treatment of influenza virus infection in Japan in February 2018. Two influenza A(H3N2) viruses carrying an I38T substitution in the polymerase acidic subunit (PA) were detected in baloxavir-treated children in December 2018. This mutation is known to confer reduced susceptibility to baloxavir, and the two mutant viruses exhibited 76- and 120-fold reduced susceptibility to baloxavir.

Influenza A(H3N2) virus exhibiting reduced susceptibility to baloxavir due to a polymerase acidic subunit I38T substitution detected from a hospitalised child without prior baloxavir treatment, Japan, January 2019.

In January 2019, two influenza A(H3N2) viruses carrying an I38T substitution in the polymerase acidic subunit (PA), which confers reduced susceptibility to baloxavir, were detected from epidemiologically unrelated hospitalised children in Japan. The viruses exhibited reduced susceptibility to baloxavir but were susceptible to neuraminidase inhibitors. Only one of the two children had been treated with baloxavir. An epidemiological analysis suggests possible transmission of the PA I38T mutant A(H3N2) virus among humans.

Complications of using Gore-Tex in medialization laryngoplasty: case series and literature review.

PURPOSE: This study was performed to evaluate the incidence and contributing factors of complications associated with medialization laryngoplasty using Gore-Tex in patients with unilateral vocal fold paralysis. METHODS: A retrospective chart review was conducted for all patients who underwent medialization laryngoplasty using Gore-Tex at Tohoku University Hospital between January 2014 and April 2018. A search of series and case reports in PubMed was performed to determine the incidence of complications following medialization laryngoplasty using Gore-Tex. RESULTS: Sixty-eight patient charts were reviewed. Two patients (2.9%) had complications (infection and extrusion into the airway) related to the Gore-Tex implant after surgery. In the 555 medialization laryngoplasty cases reported in both our current data and eight additional articles, there were 11 complications related to the Gore-Tex implant (2.0%). The most common event was extrusion into the lumen, which occurred in six cases (1.1%), followed by persistent inflammation with the granulation formation (0.5%). There were 12 cases of Gore-Tex extrusion (one male, six female, and five of unknown gender). The interval to onset ranged from 1 month to 10 years (median, 49 months). CONCLUSIONS: Our findings serve as a reminder that complications can occur with Gore-Tex implants following medialization laryngoplasty in patients with unilateral vocal fold paralysis, even in the long-term. We suggest that the use of excessively large implants in women and occurrence of postoperative hematoma followed by infection are factors that may cause complications. Nevertheless, Gore-Tex has been proven to be a relatively safe and reliable material for medialization laryngoplasty.

Poor oral intake causes enteral nutrition dependency after concomitant chemoradiotherapy for pharyngeal cancers.

PURPOSE: To identify precipitating factors responsible for enteral nutrition (EN) dependency after concomitant chemoradiotherapy (CCRT) of head and neck cancers and to examine their statistical correlations. METHODS: Factors related to feeding condition, nutritional status, disease, and treatment of 26 oropharyngeal and hypopharyngeal cancer patients who received definitive CCRT were retrospectively investigated by examining their medical records. The days of no oral intake (NOI) during hospitalization and the months using enteral nutrition after CCRT were counted as representing the feeding condition, and the changes in body weight (BW) were examined as reflecting nutritional status. The factors related to EN dependency after CCRT were analyzed. RESULTS: Long duration of total NOI (≥ 30 days) and maximum NOI ≥ 14 days were significant predictors of EN dependency. Decreased BW (≥ 7.5 kg) was the next predictor identified, but it was not significant. Multivariate analysis showed that the total duration of NOI was more correlated with EN dependency than changes in BW. CONCLUSIONS: A long duration of NOI was more strongly related to EN dependency than nutritional factors.

Influenza A(H1N1)pdm09 virus exhibiting enhanced cross-resistance to oseltamivir and peramivir due to a dual H275Y/G147R substitution, Japan, March 2016.

An influenza A(H1N1)pdm09 virus carrying a G147R substitution in combination with an H275Y substitution in the neuraminidase protein, which confers cross-resistance to oseltamivir and peramivir, was detected from an immunocompromised inpatient in Japan, March 2016. This dual H275Y/G147R mutant virus exhibited enhanced cross-resistance to both drugs compared with the single H275Y mutant virus and reduced susceptibility to zanamivir, although it showed normal inhibition by laninamivir.

Characterization of a large cluster of influenza A(H1N1)pdm09 viruses cross-resistant to oseltamivir and peramivir during the 2013-2014 influenza season in Japan.

Between September 2013 and July 2014, 2,482 influenza 2009 pandemic A(H1N1) [A(H1N1)pdm09] viruses were screened in Japan for the H275Y substitution in their neuraminidase (NA) protein, which confers cross-resistance to oseltamivir and peramivir. We found that a large cluster of the H275Y mutant virus was present prior to the main influenza season in Sapporo /: Hokkaido, with the detection rate for this mutant virus reaching 29% in this area. Phylogenetic analysis suggested the clonal expansion of a single mutant virus in Sapporo /: Hokkaido. To understand the reason for this large cluster, we examined the in vitro and in vivo properties of the mutant virus. We found that it grew well in cell culture, with growth comparable to that of the wild-type virus. The cluster virus also replicated well in the upper respiratory tract of ferrets and was transmitted efficiently between ferrets by way of respiratory droplets. Almost all recently circulating A(H1N1)pdm09 viruses, including the cluster virus, possessed two substitutions in NA, V241I and N369K, which are known to increase replication and transmission fitness. A structural analysis of NA predicted that a third substitution (N386K) in the NA of the cluster virus destabilized the mutant NA structure in the presence of the V241I and N369K substitutions. Our results suggest that the cluster virus retained viral fitness to spread among humans and, accordingly, caused the large cluster in Sapporo/Hokkaido. However, the mutant NA structure was less stable than that of the wild-type virus. Therefore, once the wild-type virus began to circulate in the community, the mutant virus could not compete and faded out.

Complete genome of hepatitis E virus from laboratory ferrets.

The complete genome of hepatitis E virus (HEV) from laboratory ferrets imported from the United States was identified. This virus shared only 82.4%-82.5% nt sequence identities with strains from the Netherlands, which indicated that the ferret HEV genome is genetically diverse. Some laboratory ferrets were contaminated with HEV.

HDR syndrome, detected in the neonatal period by newborn hearing screening.

Hypoparathyroidism, deafness, and renal dysplasia (HDR) syndrome is an autosomal dominant disorder. Because HDR syndrome is caused by haploinsufficiency in GATA3, it exhibits variation in the onset and progression of hearing loss. In previous reports, the automated auditory brainstem response (AABR) was considered insufficient to detect sensorineural hearing loss caused by HDR syndrome. We report a case of HDR syndrome whose congenital hearing loss was detected by newborn hearing screening (NHS) using AABR. In this case, HDR syndrome was suspected due to hearing loss, hypocalcemia, and her family history. Genetic testing confirmed the diagnosis of HDR syndrome at 5 months of age. Because the phenotype of hearing loss due to HDR syndrome is variable and includes progressive hearing loss, these cases may not be detected by the HNS. However, most of the previous reports were published before the NHS became common and given the frequency of hearing loss complications in HDR syndrome. We consider that there is a reasonable number of HDR syndrome cases with abnormalities on the NHS. We believe that the NHS may also be useful for early detection of hearing loss due to HDR syndrome.

The comparison of pathogenicity among SARS-CoV-2 variants in domestic cats.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has been detected or isolated from domestic cats. It is unclear whether cats play an important role in the SARS-CoV-2 transmission cycle. In this study, we examined the susceptibility of cats to SARS-CoV-2, including wild type and variants, by animal experiments. Cats inoculated with wild type, gamma, and delta variants secreted a large amount of SARS-CoV-2 for 1 week after the inoculation from nasal, oropharyngeal, and rectal routes. Only 100 TCID50 of virus could infect cats and replicate well without severe clinical symptoms. In addition, one cat inoculated with wild type showed persistent virus secretion in feces for over 28 days post-inoculation (dpi). The titer of virus-neutralizing (VN) antibodies against SARS-CoV-2 increased from 11 dpi, reaching a peak at 14 dpi. However, the omicron variant could not replicate well in cat tissues and induced a lower titer of VN antibodies. It is concluded that cats were highly susceptible to SARS-CoV-2 infection, but not to the Omicron Variant, which caused the attenuated pathogenicity.

Characterization of self-assembled virus-like particles of ferret hepatitis E virus generated by recombinant baculoviruses.

Ferret hepatitis E virus (HEV), a novel hepatitis E-like virus, has been identified in ferrets in The Netherlands. Due to the lack of a cell-culture system for ferret HEV, the antigenicity, pathogenicity and epidemiology of this virus have remained unclear. In the present study, we used a recombinant baculovirus expression system to express the 112-N-terminus and 47-C-terminus-amino-acid-truncated ferret HEV ORF2 protein in insect Tn5 cells, and found that a large amount of a 53 kDa protein (F-p53) was expressed and efficiently released into the supernatant. Electron microscopic analysis revealed that F-p53 was self-assembled into virus-like particles (ferret HEV-LPs). These ferret HEV-LPs were estimated to be 24 nm in diameter, which is similar to the size of G1, G3, G4 and rat HEV-LPs derived from both the N-terminus- and C-terminus-truncated constructs. Antigenic analysis demonstrated that ferret HEV-LPs were cross-reactive with G1, G3, G4 and rat HEVs, and rat HEV and ferret HEV showed a stronger cross-reactivity to each other than either did to human HEV genotypes. However, the antibody against ferret HEV-LPs does not neutralize G3 HEV, suggesting that the serotypes of these two HEVs are different. An ELISA for detection of anti-ferret HEV IgG and IgM antibodies was established using ferret HEV-LPs as antigen, and this assay system will be useful for monitoring ferret HEV infection in ferrets as well as other animals. In addition, analysis of ferret HEV RNA detected in ferret sera collected from a breeding colony in the USA revealed the genetic diversity of ferret HEV.

Possibility of additive effects by the presentation of visual information related to distractor sounds on the contra-sound effects of the N100m responses.

Auditory-evoked responses can be affected by different types of contralateral sounds or by attention modulation. The present study examined the additive effects of presenting visual information about contralateral sounds as distractions during dichotic listening tasks on the contralateral effects of N100m responses in the auditory-evoked cortex in 16 subjects (12 males and 4 females). In magnetoencephalography, a tone-burst of 500 ms duration at a frequency of 1000 Hz was played to the left ear at a level of 70 dB as a stimulus to elicit the N100m response, and a movie clip was used as a distractor stimulus under audio-only, visual-only, and audio-visual conditions. Subjects were instructed to pay attention to the left ear and press the response button each time they heard a tone-burst stimulus in their left ear. The results suggest that the presentation of visual information related to the contralateral sound, which worked as a distractor, significantly suppressed the amplitude of the N100m response compared with only the contralateral sound condition. In contrast, the presentation of visual information related to contralateral sound did not affect the latency of the N100m response. These results suggest that the integration of contralateral sounds and related movies may have resulted in a more perceptually loaded stimulus and reduced the intensity of attention to tone-bursts. Our findings suggest that selective attention and saliency mechanisms may have cross-modal effects on other modes of perception.

Impact of palatoplasty techniques on tympanic membrane findings and hearing prognosis in children with cleft palate.

OBJECTIVE: Children with cleft palate (CP) are at high risk of developing otitis media with effusion (OME) due to Eustachian tube (ET) dysfunction. Palatoplasty has been reported to decrease the frequency of middle ear disease and improve ET function, and although various techniques have been developed, there is no consensus on the differences in the impact of different techniques on the middle ear. The purpose of this study was to determine the differential effects of palatoplasty on middle ear function and hearing. METHODS: We performed a retrospective observational survey of pediatric patients who underwent palatoplasty for CP between June 2010 and October 2018 at Tohoku University Hospital. Cases were divided into three groups depending on the palatoplasty procedures performed: the push-back palatoplasty group, the two-flap palatoplasty group, and the Furlow double-opposing Z-plasty group. We examined the differences in clinical characteristics between patients who underwent each procedure. The primary outcome variable was tympanic membrane (TM) findings, and the secondary outcome was hearing test results. RESULTS: Children who underwent the two-flap palatoplasty had a higher tympanostomy tube (TT) insertion rate and a higher total number of TT insertions than those who underwent the Furlow double-opposing Z-plasty or the push-back palatoplasty. The TM retraction rate tended to be lower in the Furlow double-opposing Z-plasty group than in the push-back palatoplasty group or the two-flap palatoplasty group. The hearing test results at the last visit were not significantly different among the three groups. CONCLUSIONS: Children who underwent the two-flap palatoplasty had a higher rate of TT insertions, potentially increasing the risk of TM perforation. In contrast, the Furlow double-opposing Z-plasty group had a lower tendency for TM regression, possibly due to improved ET function and reduced incidence of OME. It is important to understand the advantages and disadvantages of each technique before selecting one suitable for the child's cleft and arch width. Additionally, it is important to conduct regular follow-up of TM findings and hearing test results even after palatoplasty.

Management of cholesterol granuloma due to eosinophilic otitis media using large ventilation tube.

A 46-year-old man who had been diagnosed with eosinophilic otitis media (EOM) and eosinophilic chronic rhinosinusitis was referred to our department. He suffered from bilateral earache, clogged ear sensation, and otorrhea associated with EOM. He had been treated with a myringotomy and a ventilation tube (VT) insertion. However, his symptoms did not improve, and the VT repeatedly fell out. We performed canal wall down mastoidectomy via a retro-auricular incision to remove the presumed cholesterol granuloma (CG) and a long-term VT insertion. The VT fell out repeatedly. Therefore, a large VT that Komune devised was inserted. Four months after reinsertion, there was no evidence of recurrent otorrhea or fallout of a large VT. A large VT insertion could be useful in the severe case of EOM with CG.

Use of the particle agglutination/particle agglutination inhibition test for antigenic analysis of SARS-CoV-2.

Background: The antigenicity of SARS-CoV-2 is a critical issue for the effectiveness of the vaccine, and thus, it should be phenotypically evaluated by serological assays as new field isolates emerge. The hemagglutination/hemagglutination inhibition (HA/HI) tests are well known as a representative method for antigenic analysis of influenza viruses, but SARS-CoV-2 does not agglutinate human or guinea pig red blood cells. Therefore, the antigenic analysis requires complicated cell-based assays using special equipment such as plate reader or ELISPOT analyzer. Methods: Based on the HA/HI tests for influenza viruses, we developed the particle agglutination/particle agglutination inhibition (PA/PAI) test to easily and rapidly quantify the virus and antibody using human angiotensin-converting enzyme 2 (hACE2)-bound latex beads. The virus titers were determined by mixing the beads and the virus from culture supernatant, settling it overnight, and then observing the sedimentation/agglutination pattern (PA test). The neutralization antibody titers were determined by mixing virus-infected hamster antisera in addition to the beads and virus (PAI test). Results: The PA titer was positively correlated with the plaque-forming units. The PAI titer using the hamster antisera clearly revealed the antigenic difference between the omicron and previous variants. The antigenic differences were supported by the results shown in other methods. Conclusions: The PAI test is an easy and rapid method to analyze the antigenicity of SARS-CoV-2.

Post-fusion influenza vaccine adjuvanted with SA-2 confers heterologous protection via Th1-polarized, non-neutralizing antibody responses.

Development of a universal influenza vaccine that can provide robust and long-lasting protection against heterologous infections is a global public health priority. A variety of vaccine antigens are designed to increase the antigenicity of conserved epitopes to elicit cross-protective antibodies that often lack virus-neutralizing activity. Given the contribution of antibody effector functions to cross-protection, adjuvants need to be added to modulate antibody effector functions as well as to enhance antibody quantity. We previously showed that post-fusion influenza vaccine antigens elicit non-neutralizing but cross-protective antibodies against conserved epitopes. Here, using a murine model, we comparably assessed the adjuvanticity of the newly developed SA-2 adjuvant containing a synthetic TLR7 agonist DSP-0546 and squalene-based MF59 analog as representative Th1- or Th2-type adjuvants, respectively. Both types of adjuvants in the post-fusion vaccine comparably enhanced cross-reactive IgG titers against heterologous strains. However, only SA-2 skewed the IgG subclass into the IgG2c subclass in association to its Th1-polarizing nature. SA-2-enhanced IgG2c responses exhibited antibody-dependent cellular cytotoxicity against heterologous virus strains, without cross-neutralizing activity. Eventually, the SA-2-adjuvanted vaccination provided protection against lethal infection by heterologous H3N2 and H1N1 viruses. Together, we conclude that the combination with a SA-2 is advantageous for enhancing the cross-protective capability of post-fusion HA vaccines that elicit non-neutralizing IgG antibodies.