Epidemiological and clinical characteristics of infections with seasonal human coronavirus and respiratory syncytial virus in hospitalized children immediately before the coronavirus disease 2019 pandemic.

INTRODUCTION: Seasonal human coronavirus (HCoV)-229E, -NL63, -OC43, and -HKU1 are seasonal coronaviruses that cause colds in humans. However, the clinical characteristics of pediatric inpatients infected with HCoVs are unclear. This study aimed to compare and clarify the epidemiological and clinical features of HCoVs and respiratory syncytial virus (RSV), which commonly causes severe respiratory infections in children. METHODS: Nasopharyngeal swabs were collected from all pediatric inpatients with respiratory symptoms at two secondary medical institutions in Fukushima, Japan. Eighteen respiratory viruses, including RSV and four HCoVs, were detected via reverse transcription-polymerase chain reaction. RESULTS: Of the 1757 specimens tested, viruses were detected in 1272 specimens (72.4%), with 789 single (44.9%) and 483 multiple virus detections (27.5%). RSV was detected in 639 patients (36.4%) with no difference in clinical characteristics between RSV-A and RSV-B. HCoV was detected in 84 patients (4.7%): OC43, NL63, HKU1, and 229E in 25 (1.4%), 26 (1.5%), 23 (1.3%), and 16 patients (0.9%), respectively. Patients with HCoV monoinfection (n = 35) had a significantly shorter period from onset to hospitalization (median [interquartile range] days, 2 [1-4.5] vs. 4 [2-5]), significantly shorter hospitalization stays (4 [3-5] vs. 5 [4-6]), and more cases of upper respiratory infections (37.1% vs. 3.9%) and croup (17.1% vs. 0.3%) but less cases of lower respiratory infection (54.3% vs. 94.8%) than patients with RSV monoinfection (n = 362). CONCLUSION: Seasonal HCoV-infected patients account for approximately 5% of children hospitalized for respiratory tract infections and have fewer lower respiratory infections and shorter hospital stays than RSV-infected patients.

Detection of Variants With Reduced Baloxavir Marboxil and Oseltamivir Susceptibility in Children With Influenza A During the 2019-2020 Influenza Season.

BACKGROUND: We aimed to detect influenza variants with reduced susceptibility to baloxavir marboxil (baloxavir) and oseltamivir and identify differences in the clinical course between children with and without these variants after antiviral treatment. METHODS: During the 2019-2020 influenza season, we enrolled children with confirmed influenza A (20 treated with baloxavir and 16 with oseltamivir). We analyzed patients' sequential viral RNA loads and infectious virus titers, the drug susceptibilities of clinical isolates, and amino acid substitutions in the viral polymerase acidic protein subunits or neuraminidase. We assessed patients' clinical information using questionnaires. RESULTS: All viral RNA loads and virus titers were significantly decreased after treatment, but we detected baloxavir-resistant and oseltamivir-resistant variants in 5 of 20 and 3 of 16 patients, respectively. The duration of fever was similar between patients with and without the variants, but infectious viral shedding lasted 3 days longer in patients with baloxavir-resistant variants. In addition, the duration to improvement of clinical symptoms was longer in these patients (75.0 vs 29.5 hours; P = .106). CONCLUSIONS: After antiviral treatment, the emergence of baloxavir-resistant variants may affect the patients' clinical course, but oseltamivir-resistant variants had no clinical impact.

Towards the development of a human in vitro model of the blood-brain barrier for virus-associated acute encephalopathy: assessment of the time- and concentration-dependent effects of TNF-α on paracellular tightness.

The pathogenesis of virus-associated acute encephalopathy (VAE) involves brain edema caused by disruption of the blood-brain barrier (BBB). We aimed to develop an in vitro VAE model using an in vitro BBB model, to evaluate the dynamics of vascular dysfunction caused by tumor necrosis factor (TNF)-α. A co-culture model, consisting of Transwell®-grown human brain microvascular endothelial cells and pericytes, was treated with serially diluted TNF-α. Transendothelial electrical resistance (TER) was measured using cellZscope®. A permeability assay, using fluorescein isothiocyanate-conjugated sodium or dextran, was performed. Changes in claudin-5 localization and expression after TNF-α treatment were observed using immunofluorescence staining and western blot analysis. The TER decreased and permeability increased after TNF-α treatment; recovery time was dependent on TNF-α concentration. Claudin-5 was delocalized after TNF-α treatment and recovered in a TNF-α concentration-dependent manner. The expression of claudin-5 decreased 24 h after the TNF-α treatment and completely recovered 48 h after TNF-α treatment. Claudin-5 delocalization was likely associated with vascular hyperpermeability. To conclude, we evaluated vascular endothelial cell permeability and injury in VAE using an in vitro BBB model treated with TNF-α. This system can be useful for developing novel therapeutic strategies for VAE and designing treatments that target vascular permeability.

Detection of Variants With Reduced Baloxavir Marboxil Susceptibility After Treatment of Children With Influenza A During the 2018-2019 Influenza Season.

During the 2018-2019 influenza seasons, we detected reduced baloxavir marboxil (baloxavir) susceptible variants with I38S or I38T amino acid substitutions on the PA subunit of influenza virus ribonucleic acid polymerase in 7 of 18 baloxavi-treated children and found that virus titer rebounded in some of these children with variants. We also found fever durations to be similar between patients with or without the variants, but the patients with variants shed the virus 3 days longer and took longer to improve clinical symptoms than those without variants. The emergence of these variants should be monitored during future influenza seasons.

Utility of enzyme immunoassays for diagnosis of subacute sclerosing panencephalitis.

BACKGROUND: Subacute sclerosing panencephalitis (SSPE) is a progressive neurologic disorder caused by the measles virus (MV) and is identified by positive MV-specific antibody titers, detected mainly by hemagglutination inhibition (HI) tests in the cerebrospinal fluid (CSF). However, an alternative method, the enzyme immunoassay (EIA), has increasingly become a preferred method for detecting MV antibodies. To establish the index for SSPE diagnosis using EIA, we investigated the correlation between HI and EIA titers of MV antibodies in SSPE patients. METHODS: Data on MV antibody titers and measurement methods at the time of diagnosis in 89 Japanese SSPE cases diagnosed between 1979 and 2006 were obtained by a survey. We also assessed the serum and CSF MV antibody titers in three patients with SSPE and serum MV antibody titers in 38 healthy adults using immunoglobulin G (IgG)-EIA and HI. RESULTS: In all cases diagnosed as SSPE, IgG-EIA titers in the CSF were ≥0.49 IU/mL. There was a positive correlation between serum antibody values in the controls measured by IgG-EIA and HI. In patients with SSPE, both serum and CSF antibody values, measured by IgG-EIA, and HI, were positively correlated, and a positive correlation was found between the serum and CSF MV antibody titers as measured by IgG-EIA. The serum/CSF MV antibody titer ratios determined by IgG-EIA were <20 in most SSPE patients. CONCLUSIONS: Immunoglobulin G-EIA may be a suitable alternative method for SSPE diagnosis; however, its potential utility and the cut-off point of ≥0.49 IU/mL should be tested with additional patient cohorts.

Case report of severe myocarditis in an immunocompromised child with Respiratory Syncytial Virus infection.

BACKGROUND: Respiratory syncytial virus (RSV) infection is common and may be severe among patients with preexisting cardiac anomalies, but direct involvement of myocardial damage is not common in those patients. Additionally, myocardial involvement has been rarely described among immune compromised children. CASE PRESENTATION: A 4-year-old girl with acute lymphoblastic leukemia who received maintenance chemotherapy in an outpatient clinic developed systemic inflammatory response syndrome. RSV infection was confirmed by a positive rapid antigen test and serological assay. Subsequently, she was diagnosed with severe myocarditis caused by RSV infection, which was diagnosed by abnormal findings of cardiac echography and ECG and elevated biomarkers for myocardial damage. Then, she was treated in the intensive care unit for 13 days. High amounts of RSV type B RNA was detected in tracheal aspirates and serum sample. CONCLUSION: This case report emphasizes that RSV infection may be associated with myocarditis in immunocompromised children receiving maintenance chemotherapy.

Establishment of a method for evaluating endothelial cell injury by TNF-α in vitro for clarifying the pathophysiology of virus-associated acute encephalopathy.

BACKGROUND: Virus-associated acute encephalopathy (VAE) is a severe central nervous system complication caused by common viral infections in children. The pathophysiology of VAE is thought to be endothelial injury. This study was designed to establish an in vitro VAE model for evaluating endothelial injury caused by the proinflammatory cytokine TNF-α. METHODS: Transwell-grown human umbilical vein endothelial cells (HUVECs) monolayers were incubated with serially diluted TNF-α. Transendothelial electrical resistance (TER) was measured using impedance spectroscopy. Permeability changes of HUVECs after TNF-α treatment were determined by fluorescein isothiocyanate (FITC)-conjugated dextran. Moreover, TNF-α-induced morphological changes in claudin-5 and apoptosis were observed by immunofluorescent staining. RESULTS: The decrease in TER, time of TER recovery to baseline, and increase in permeability were all dependent on TNF-α concentration. Immunofluorescent staining showed that claudin-5 was delocalized after TNF-α treatment in a dose-dependent manner. In addition, some apoptotic cells were observed at high TNF-α concentrations. CONCLUSION: TER measurement combined with a permeability assay could be useful for evaluating vascular endothelial cell permeability in an in vitro model. These evaluation methods will contribute to both the development of specific treatments focusing on vascular permeability, and the search for a novel therapeutic strategy in VAE treatment.

Respiratory syncytial virus shedding by children hospitalized with lower respiratory tract infection.

Children with respiratory syncytial virus (RSV) infection shed virus for variable periods. The aim of this study was to quantify the viral load in nasopharyngeal aspirates of children with RSV throughout their hospitalization. This study included 37 children who were admitted with a diagnosis of RSV infection based on a positive rapid diagnostic test. Nasopharyngeal aspirates were collected from patients every day, from admission to discharge. Viral detection and quantification were performed using quantitative real-time PCR. Of the 37 patients, RSV-A was detected in 29 and RSV-B in 6. Two patients were PCR-negative for any type of RSV. RSV-A was detected in 12 of 16 patients (75%) 6 days after admission. These patients shed detectable virus from days 1 to 12, and for a significantly longer period (mean 5.7 days) than RSV-B (mean 3.8 days) patients. Half of the RSV-A patients were also positive on day 14 following onset. RSV-A was detected in patients <12 months of age for significantly longer periods after onset than in patients ≥12 months of age. RSV-A viral load was negatively correlated with days from admission and days from onset. Because RSV shedding was frequently prolonged, the hospitalized children may have contracted RSV as a nosocomial infection. To prevent nosocomial RSV infections in hospital wards, healthcare workers must take appropriate infection control measures and provide adequate guidance on hand washing to the family of the patient.

Differences in clinical findings, pathology, and outcomes between C3 glomerulonephritis and membranoproliferative glomerulonephritis.

BACKGROUND: To clarify the clinical manifestations of pediatric complement component C3 glomerulonephritis (C3GN), we retrospectively evaluated differences in the clinicopathological findings and prognosis between C3GN and immune-complex-mediated membranoproliferative glomerulonephritis (IC-MPGN). METHODS: Thirty-seven patients diagnosed with "idiopathic MPGN" were enrolled in this retrospective study. The patients were divided into two groups, with Group 1 consisting of 19 patients diagnosed with IC-MPGN and Group 2 consisting of 18 patients diagnosed with C3GN. The clinical findings and the prognosis were investigated for both groups. RESULTS: Thirteen patients in Group 2 were identified by mandatory annual school screening for urinary abnormalities. The incidence of macro-hematuria and the frequency of low serum C4 values were lower in Group 2 patients than in Group 1 patients. At the time of the second renal biopsy, urinary protein excretion, incidence of hematuria, frequency of low serum C3 values, and scores for mesangial proliferation, glomerular sclerosis, and interstitial fibrosis were higher in Group 2 patients than in Group 1 patients. At the most recent follow-up examination, the number of patients categorized as non-responding or with end-stage renal disease was higher in Group 2 patients than in Group 1 patients. CONCLUSIONS: Our results suggest that the treatment response and prognosis of patients with C3GN are worse than those of patients with IC-mediated MPGN. Therefore, in the clinical context regarding treatment options and prognosis, it may be useful to classify idiopathic MPGN as C3GN or IC-MPGN. In addition, long-term follow-up of C3GN is necessary.

Influenza viral load and peramivir kinetics after single administration and proposal of regimens for peramivir administration against resistant variants.

We estimated the efficacy of the current single administration of peramivir on the basis of peramivir pharmacokinetics in the upper respiratory tract (URT) and determined the predictive peramivir concentration-time curve to assess its efficacy against viruses with decreased susceptibility to neuraminidase inhibitors. Serum, nasal swab, or aspiration samples were collected from 28 patients treated with 10 mg/kg body weight peramivir. The sequential influenza viral RNA load and susceptibility after peramivir administration were measured using a quantitative real-time reverse transcription-PCR and neuraminidase inhibition assay. The peramivir concentrations in the serum and URT after a single administration at 10 mg/kg were measured, and the predictive blood and URT peramivir concentration-time curves were determined to assess various administration regimens against resistant variants. The peramivir concentration decreased to <0.1% of the maximum concentration of drug in serum (Cmax) at 24 h after administration. Rapid elimination of peramivir from the URT by 48 h after administration may contribute to an increase in the influenza A viral load after day 3 but not to a decrease in the influenza B viral load, despite the absence of a decrease in the susceptibility to peramivir. A longer maintenance of a high level of peramivir in the URT is expected by divided administration rather than once-daily administration. When no clinical improvement is observed in patients with normal susceptibility influenza A and B, peramivir readministration should be considered. In severe cases caused by resistant variants, better inhibitory effectiveness and less frequent adverse events are expected by divided administration rather than once-daily administration with an increased dosage.

The efficacy of recombinant human soluble thrombomodulin for the treatment of shiga toxin-associated hemolytic uremic syndrome model mice.

BACKGROUND: Recombinant human soluble thrombomodulin (rhTM) is a promising therapeutic natural anticoagulant that is comparable to antithrombin, tissue factor pathway inhibitor and activated protein C. In order to clarify the efficacy of rhTM for the treatment of typical hemolytic uremic syndrome (t-HUS), we examined changes in renal damage in t-HUS mice treated with rhTM or vehicle alone. METHODS: We used severe and moderate t-HUS mice injected with shiga toxin (Stx) and lipopolysaccharide (LPS). The severe t-HUS mice were divided into two subgroups [an rhTM subgroup (Group A) and a saline subgroup (Group B)] along with the moderate t-HUS mice [an rhTM subgroup (Group C) and a saline subgroup (Group D)]. Groups E and F were healthy mice treated with rhTM or saline, respectively. RESULTS: All mice in Group B died at 80-90 h post-administration of Stx2 and LPS whereas all mice in Group A remained alive. Loss of body weight, serum creatinine level, endothelial injury and mesangiolysis scores at 24 h after administration in the t-HUS mice treated with rhTM were lower than those in t-HUS mice treated with saline. The levels of hemoglobin at 6 h and platelet counts at 24 h after administration in Group A were higher than those in Group B. Serum interleukin (IL)-6, IL-1ß and tumor necrotic factor (TNF)-α levels at 24 h after administration in Group A were lower than those in Group B. Serum C5b-9 levels at 24 h after the administration and serum fibrinogen degradation product (FDP) at 72 h after the administration of Stx2 and LPS were lower in Group A than in Group B. CONCLUSIONS: These results indicate that rhTM might afford an efficacious treatment for t-HUS model mice via the inhibition of further thrombin formation and amelioration of hypercoagulant status.

Influenza A H1N1 pdm09-associated myocarditis during zanamivir therapy.

A 9-year-old girl developed influenza A H1N1 pdm09-associated myocarditis and pericarditis 2 days after starting zanamivir therapy. The virus was detected in the respiratory tract but not in the serum or pericardial effusion. The virus sampled from the respiratory tract had normal susceptibility to neuraminidase inhibitors. Although no differences in interferon-γ, interleukin (IL)-1ß, and tumor necrosis factor-α were observed between the plasma and pericardial effusion, some inflammatory cytokines or chemokines (IL-6 and IL-8) and vascular endothelial growth factor were remarkably elevated in the pericardial effusion compared with the plasma. This suggested that the influenza virus, after infecting the respiratory tract, affected the myocardium, causing myocarditis to gradually develop, which might have been followed by an autoreactive pericarditis causing increased pericardial effusion. Therefore, influenza-associated myocarditis should be considered when influenza patients have respiratory and cardiac involvement, even during treatment with a neuraminidase inhibitor.

Incidence and prognosis of systemic lupus erythematosus in a 35 year period in Fukushima, Japan.

BACKGROUND: We examined the epidemiology, clinical manifestations, and prognosis of pediatric systemic lupus erythematosus (SLE) in Fukushima Prefecture, Japan over a 35 year period. METHODS: We collected the medical records of 37 patients diagnosed with SLE between 1977 and 2013. These children were divided into two groups. group 1 consisted of 19 patients who were diagnosed between 1977 and 1995, and group 2 consisted of 18 patients diagnosed between 1996 and 2013. The epidemiology, clinical features, and prognosis were retrospectively compared between the two groups. RESULTS: The mean number of patients per 100,000 children per year for group 1 and group 2 was 0.33 ± 0.25 and 0.35 ± 0.30, respectively. The duration from onset of symptoms to treatment in group 2 was shorter than that in group 1, but the clinical and laboratory findings at onset did not differ between the two groups. All patients were treated with prednisolone, and 17 patients in group 1 and 18 in group 2 were treated with methylprednisolone pulse therapy. The frequency of cyclophosphamide treatment decreased whereas the frequency of cyclosporine, tacrolimus and mizoribine pulse therapy increased in group 2. SLE disease activity index (SLEDAI) score at the latest follow up in group 2 was lower in group 1. The survival rate was 84% in group 1 and 100% in group 2. CONCLUSION: The frequency and severity of SLE in group 1 were similar to those in group 2, and the prognosis of SLE in group 2 was better than that in group 1.

Characteristics of viruses derived from nude mice with persistent measles virus infection.

Measles virus (MV) isolates from patients with subacute sclerosing panencephalitis (SSPE) differ from wild-type MV virologically. However, few animal models have reported viruses with characteristics of the SSPE virus. The MV Edmonston strain was inoculated into the subarachnoid space of nude mice. All nude mice displayed weight loss and required euthanasia, with a mean survival duration of 73.2 days. The viral load in the brain was 4- to 400-fold higher than the inoculated load, and brain infection was confirmed by immunostaining. Gene sequencing of the viruses revealed that amino acid mutations occurred more frequently in matrix proteins. The most common mutation was a uridine-to-cytosine transition. The virus exhibited lower free virus particle formation ability than the Edmonston strain. When nude mice were challenged with 2 × 10(2) PFU of the brain-derived virus, the mean survival duration was 34.7 days, which was significantly shorter than that of the mice challenged with 4 × 10(4) PFU of the Edmonston strain (P < 0.01). This study indicated that MV in a nude mouse model of persistent infection exhibited characteristics of the SSPE virus. This model may prove useful in elucidating the pathogenic mechanism of SSPE and developing potential therapeutics.

Clinical and epidemiologic factors related to subsequent wheezing after virus-induced lower respiratory tract infections in hospitalized pediatric patients younger than 3 years.

UNLABELLED: Most wheezing episodes in infants are caused and exacerbated by virus-induced lower respiratory tract infections. However, there are few reports of epidemiologic and clinical virus-specific research with a focus on virus-induced wheezing. The purpose of the current study was to characterize the clinical presentation of virus-induced wheezing in pediatric patients <3 years of age who were hospitalized with lower respiratory tract infections. Of the 412 patients in the study, 216 were followed for 3 years. Nasopharyngeal aspirates collected from the patients at the time of admission were examined for the presence of respiratory syncytial virus (RSV), rhinovirus (RV), parainfluenza-3 virus (PIV-3), human metapneumovirus (hMPV), and influenza virus (Flu) using reverse-transcription polymerase chain reaction and rapid diagnostic tests. Clinical signs were assessed using a severity scoring system. In patients with wheezing at the time of admission, RSV, RV, RSV+RV, Flu, PIV-3, and hMPV were detected in 33, 14, 8, 8, 5, and 3 % of samples, respectively. There were no differences in age and severity scores between patients harboring more prevalent viruses (RSV and RV) and those with less common infections. Patients with wheezing and RV-positive aspirates at the time of admission were more likely to develop subsequent wheezing during the following 3 years. CONCLUSION: RSV and RV infections are factors in the development and exacerbation of wheezing after virus-induced lower respiratory tract infections. Moreover, RV-induced wheezing may be associated with subsequent recurrent wheezing and the development of asthma.

Resistance factors for the treatment of immunoglobulin A nephropathy with diffuse mesangial proliferation.

AIM: Some patients with severe immunoglobulin A nephropathy (IgAN) are resistant to multi-drug combination therapy; however, there have been few reports on the risk factors for non-responsiveness to treatment for severe IgAN. We, therefore, evaluated the risk factors for non-responsiveness to treatment in cases of severe IgAN. METHODS: We collected data on 44 children who had been diagnosed with IgAN with diffuse mesangial proliferation and treated with multi-drug combination therapy. The children were divided into two groups based on the prognosis at the latest follow-up. Group 1 consisted of 30 children with normal urine and nine children with minor urinary abnormalities and Group 2 consisted of four children with persistent nephropathy and one child with renal insufficiency. The clinical, laboratory, and pathological findings for both groups were analyzed. RESULTS: The age at the onset in Group 2 was higher than that in Group 1. C3 deposits and high chronicity index values at the first renal biopsy were more frequently found in Group 2 than in Group 1 patients. IgA deposits, serum IgA and myeloid-related protein (MRP) 8/14 levels, and glomerular and interstitial MRP8+CD68+ scores at the second biopsy were all higher in Group 2 than in Group 1 patients. CONCLUSIONS: Our results, although based on only a small number of patients in a retrospective study, suggest that age, presence of C3 deposits and interstitial changes at the onset, and persistent renal inflammatory activation may be risk factors for non-responsiveness to treatment for IgAN with diffuse mesangial proliferation.

Molecular Epidemiology of Human Metapneumovirus in East Japan before and after COVID-19, 2017-2022.

Human metapneumovirus (hMPV) is genetically classified into two major subgroups, A and B, based on attachment glycoprotein (G protein) gene sequences. The A2 subgroup is further separated into three subdivisions, A2a, A2b (A2b1), and A2c (A2b2). Subgroup A2c viruses carrying 180- or 111-nucleotide duplications in the G gene (A2c 180nt-dup or A2c 111nt-dup ) have been reported in Japan and Spain. The coronavirus disease 2019 (COVID-19) pandemic disrupted the epidemiological kinetics of other respiratory viruses, including hMPV. In this study, we analyzed the sequences of hMPV isolates in Tokyo and Fukushima obtained from 2017 to 2022, i.e., before and after the COVID-19 pandemic. Subgroup A hMPV strains were detected from 2017 to 2019, and most cases were A2c 111nt-dup, suggesting ongoing transmission of this clade, consistent with global transmission dynamics. Subgroup B viruses, but not subgroup A viruses, were detected in 2022 after the COVID-19 peak. Phylogenetic analysis showed that the subgroup B viruses were closely related to strains detected in Yokohama from 2013 to 2016, and strains detected in Fukushima in 2019, suggesting the reappearance of local endemic viruses in East Japan.

Treatment with oseltamivir and prone positioning for a 9-year-old boy with ARDS caused by pandemic H1N1 2009 influenza.

Here we report the case of a 9-year-old boy with acute respiratory distress syndrome (ARDS) caused by novel H1N1 swine-origin influenza virus A. A diagnosis of ARDS caused by a novel influenza A (H1N1) virus was made on the basis of chest X-ray and computed tomography together with low oxygenation index (OI) and the detection of novel influenza A (H1N1) virus from tracheal secretion samples. Oseltamivir phosphate and prone positioning were effective in the treatment of ARDS in this case. These findings suggest that anti-viral drugs and prone positioning can play an important role in the improvement of ARDS caused by novel H1N1 swine-origin influenza virus A.

Clinical and Virologic Impacts of Respiratory Viral Co-infections in Children With Influenza.

BACKGROUND: Advances in multiplex polymerase chain reaction (PCR) methods have enabled the simultaneous detection of multiple respiratory viruses. We aimed to estimate the clinical and virologic impacts of influenza and other respiratory virus co-infection in children. METHODS: We enrolled 38 and 35 children diagnosed with influenza and treated with baloxavir marboxil (baloxavir) and oseltamivir, respectively. We performed quantitative reverse transcription-PCR to detect and measure the levels of noninfluenza viruses from 3 nasopharyngeal swab samples collected before and on days 3 and 5 after the initial antiviral dose. We assessed patients' clinical information using questionnaires. RESULTS: One or more respiratory viruses other than influenza virus were detected in 26 (35.6%) of 73 children before antiviral treatment. The influenza virus load and clinical characteristics on the day of influenza onset were similar between children with and without virus co-infections. Of the 26 and 32 children without the emergence of the reduced baloxavir and oseltamivir susceptible variants after treatment, 8 (30.8%) and 7 (21.9%) children were dually co-infected with human rhinovirus only, respectively. The level of human rhinovirus RNA on day 0 in these children was less than -3 log 10 that of influenza virus RNA, and the human rhinovirus co-infection had no impact on the disease course either clinically or virologically. CONCLUSIONS: When multiple respiratory viruses are detected in the same patient, it is necessary to assess clinical symptoms as well as the levels of detected viruses to determine which virus contributes to the development of illness.

Serotypes and Antibiotic Resistance of Streptococcus pneumoniae before and after the Introduction of the 13-Valent Pneumococcal Conjugate Vaccine for Adults and Children in a Rural Area in Japan.

The increase in non-vaccine serotypes of Streptococcus pneumoniae and their multidrug resistance have become an issue following the introduction of the 13-valent pneumococcal conjugate vaccine (PCV13). In this study, we investigated the serotypes and drug resistance of S. pneumoniae detected in adult and pediatric outpatients at a hospital in a rural area of Japan between April 2012 and December 2016. Serotypes of the bacterium were identified using the capsular swelling test and multiplex polymerase chain reaction testing of DNA extracted from the specimens. Antimicrobial susceptibility was determined using the broth microdilution method. The serotype 15A was classified using multilocus sequence typing. The results showed that the prevalence of non-vaccine serotypes increased significantly in children from 50.0% in 2012-2013 to 74.1% in 2016 (p ≤ 0.006) and in adults from 15.8% in 2012-2013 to 61.5% in 2016 (p ≤ 0.026), but no increase in drug-resistant isolates was evident. However, an increase in the drug-resistant serotypes 15A and 35B was observed in children. Although isolates of these two serotypes showed cefotaxime susceptibility, cefotaxime resistance was confirmed for the serotype 15A isolates. Future trends in the spread of these isolates should be monitored with caution.