Outcomes of hematopoietic stem cell gene therapy for Wiskott-Aldrich syndrome.

Wiskott-Aldrich syndrome (WAS) is a rare X-linked disorder characterized by combined immunodeficiency, eczema, microthrombocytopenia, autoimmunity, and lymphoid malignancies. Gene therapy (GT) to modify autologous CD34+ cells is an emerging alternative treatment with advantages over standard allogeneic hematopoietic stem cell transplantation for patients who lack well-matched donors, avoiding graft-versus-host-disease. We report the outcomes of a phase 1/2 clinical trial in which 5 patients with severe WAS underwent GT using a self-inactivating lentiviral vector expressing the human WAS complementary DNA under the control of a 1.6-kB fragment of the autologous promoter after busulfan and fludarabine conditioning. All patients were alive and well with sustained multilineage vector gene marking (median follow-up: 7.6 years). Clinical improvement of eczema, infections, and bleeding diathesis was universal. Immune function was consistently improved despite subphysiologic levels of transgenic WAS protein expression. Improvements in platelet count and cytoskeletal function in myeloid cells were most prominent in patients with high vector copy number in the transduced product. Two patients with a history of autoimmunity had flares of autoimmunity after GT, despite similar percentages of WAS protein-expressing cells and gene marking to those without autoimmunity. Patients with flares of autoimmunity demonstrated poor numerical recovery of T cells and regulatory T cells (Tregs), interleukin-10-producing regulatory B cells (Bregs), and transitional B cells. Thus, recovery of the Breg compartment, along with Tregs appears to be protective against development of autoimmunity after GT. These results indicate that clinical and laboratory manifestations of WAS are improved with GT with an acceptable safety profile. This trial is registered at clinicaltrials.gov as #NCT01410825.

Molecular Evolution and Epidemiology of Parechovirus-A3 in Japan, 1997-2019.

Parechovirus-A3 (PeV-A3), first reported in 2004 in Japan, is an emerging pathogen that causes sepsis and meningoencephalitis in neonates and young infants. Although PeV-A3 has been identified worldwide, its epidemiological characteristics differ by region. To investigate the molecular evolution and epidemiology of PeV-A3, we performed genetic analyses of 131 PeV-A3 strains from the years 1997-2019 in Niigata, Japan. During 2016-2019, annual numbers remained steady, in contrast to the PeV-A3 epidemic interval of every 2-3 years that was observed in Japan from 2006. Bayesian evolutionary analysis of the complete viral protein 1 region revealed alternate dominant clusters during years of PeV-A3 epidemics. The branch including the oldest and first isolated PeV-A3 strains in Japan has been disrupted since 2001. The year of PeV-A3 emergence was estimated to be 1991. Continuous surveillance with genetic analyses of different regions will improve understanding of PeV-A3 epidemiology worldwide.

Detection of parechovirus-A in hospitalized children with acute lower respiratory infection in Myanmar, 2017-2018.

Parechovirus-A (PeV-A) causes emerging infection in children, and clinical presentation depends on genotype. The virus has been investigated mainly in developed countries; however, data from developing countries, especially in Asia, are sparse. This study investigated whether PeV-A circulated in children in Myanmar. This retrospective study evaluated PeV-A in nasopharyngeal samples from children aged 1 month to 12 years who were hospitalized with acute lower respiratory infection at Yankin Children Hospital, Yangon, Myanmar, during the period from May 2017 to April 2019. Real-time polymerase chain reaction (PCR) was used to detect PeV-A, and PCR-positive samples were used for genotyping and phylogenetic analysis. In total, 11/570 (1.9%) of samples were positive for PeV-A; 7 were successfully genotyped by sequencing the VP3/VP1 region, as follows: PeV-A1 (n = 4), PeV-A5 (n = 1), PeV-A6 (n = 1), and PeV-A14 (n = 1). Median age was 10.0 months (interquartile range 4.0-12.0 months), and other respiratory viruses were detected in all cases. Phylogenetic analysis showed that all detected PeV-A1 strains were in clade 1 A, which was a minor clade worldwide. Four PeV-A genotypes were detected in Myanmar. The clinical impact of PeV-A in children should be evaluated in future studies.

Purulent nasal discharge due to gonococcal nasopharyngitis in a neonate.

Neonatal gonorrhea, caused by Neisseria gonorrhoeae, is an uncommon but important disease to prevent because its complications, such as gonococcal ophthalmia neonatorum causes blindness if untreated. Neonatal gonococcal nasopharyngitis is a rare, but important clinical manifestation to suspect gonococcal infection in a neonate. Herein we report a case of neonatal gonococcal nasopharyngitis, presented with purulent nasal discharge. A full-term male neonate without perinatal complications developed purulent eye discharge on the 7th day of life. N. gonorrhoeae was isolated from the eye discharge culture; however, he did not receive the standard regimen. Subsequently, he presented to our hospital with fever and nasal discharge on the 20th day of life. N. gonorrhoeae was also isolated from nasal discharge and nasopharyngeal swabs without any evidence of chlamydia or syphilis. He received intravenous cefotaxime until disseminated gonococcal infection was ruled out and was discharged without any sequelae. Rhinorrhea in newborns requires consideration of mother-to-child transmission of various microorganisms, not only common respiratory viruses, but also rare, serious preventable infections such as gonorrhea or syphilis. Along with the recent syphilis patients on the rise in Japan, gonorrhea is an important disease to recognize, and the incidence could increase. Clinical manifestations of neonatal gonococcal infections, including nasopharyngitis, need to be recognized to suspect the diagnosis and initiate appropriate treatment to prevent serious complications.

Multiple pneumatoceles and lung abscesses caused by early-onset E. coli sepsis in a preterm neonate.

Early-onset sepsis (EOS) is a serious and fatal illness in neonates, Group B Streptococcus and Escherichia coli are major causative pathogens. We report a case of EOS and pneumonia caused by E. coli in a preterm neonate with multiple pneumatoceles and lung abscesses. A male neonate weighing 1670g was delivered at 33 6/7 weeks' gestation by a mother with clinical chorioamnionitis. He showed respiratory distress soon after birth and developed septic shock. He was intubated and mechanical ventilation was started. E.coli was detected in blood culture obtained from both the patient and his mother. He developed multiple pneumatoceles and lung abscesses. Surgical drainage was complicated, cefotaxime was thus continued until day 74. Pneumatoceles and lung abscesses are complications of neonatal pneumonia, rarely reported by E. coli. Multiple lung abscesses in our patient are distinct from single abscesses in previous case studies of neonatal lung abscesses. We speculate that bacteremia along with pneumatoceles led to multiple lung abscesses in our patient. These complications require long-term antibiotic therapy, to minimize morbidity and mortality, and should thus be considered when managing EOS caused by E. coli.

Clinical manifestations and outcome of viral acute lower respiratory infection in hospitalised children in Myanmar.

BACKGROUND: Acute lower respiratory infection (ALRI) remains the leading cause of death in children worldwide, and viruses have been the major cause of ALRI. In Myanmar, ALRI is associated with high morbidity and mortality in children, and detailed information on ALRI is currently lacking. METHODS: This prospective study investigated the viral aetiologies, clinical manifestations, and outcomes of ALRI in hospitalised children aged 1 month to 12 years at the Yankin Children Hospital, Yangon, Myanmar from May 2017 to April 2019. The sample size was set to 300 patients for each year. Two nasopharyngeal swabs were obtained for the patients with suspected viral ALRI; one for rapid tests for influenza and respiratory syncytial virus (RSV), and the other for real-time PCR for the 16 ALRI-causing viruses. Pneumococcal colonization rates were also investigated using real-time PCR. Clinical information was extracted from the medical records, and enrolled patients were categorised by age and severity for comparison. RESULTS: Among the 5463 patients admitted with a diagnosis of ALRI, 570 (10.4%) were enrolled in this study. The median age of the patients was 8 months (interquartile range, 4-15 months). The most common symptoms were cough (93%) and difficulty in breathing (73%), while the most common signs of ALRI were tachypnoea (78%) and chest indrawing (67%). A total of 16 viruses were detected in 502 of 570 patients' samples (88%), with RSV B (36%) and rhinovirus (28%) being the most commonly detected. Multiple viruses were detected in 221 of 570 samples (37%) collected from 570 patients. Severe ALRI was diagnosed in 107 of 570 patients (19%), and RSV B and human rhinovirus were commonly detected. The mortality rate was 5%; influenza virus A (29%) and RSV B (21%) were commonly detected, and stunting and lack of immunization were frequently observed in such cases. Additionally, 45% (259/570) of the patients had pneumococcal colonization. CONCLUSIONS: Viral ALRI in hospitalised children with a median of 8 months has significant morbidity and mortality rates in Myanmar. RSV and rhinovirus were the most commonly detected from nasopharyngeal swabs, while influenza virus and RSV were the most frequently associated with fatal cases.

An atypical case of Kawasaki disease with severe pneumonia in a neonate.

BACKGROUND: Kawasaki disease (KD) is an acute, febrile, systemic vasculitis of unknown etiology that primarily affects the coronary arteries and generally occurs at around 1 year of age. Although the diagnosis of KD is generally not difficult, it is challenging in cases of incomplete KD lacking characteristic clinical manifestations. The incidence of incomplete KD is higher in infants younger than 6 months of age. Pneumonia is an extremely rare complication of KD and can be misinterpreted as atypical pneumonia rather than KD. Herein, we report a neonate with atypical KD and severe pneumonia who required mechanical ventilation. CASE PRESENTATION: Japanese one-month-old infant had only fever and rash on admission (day 1), and he was transferred to the intensive care unit for severe pneumonia on day 2. Although pneumonia improved following intensive care, he was diagnosed with KD on day 14 because of emerging typical clinical manifestations such as fever, bulbar nonexudative conjunctival injection, desquamation of the fingers, and coronary artery aneurysm. KD symptoms improved after three doses of intravenous immunoglobulin plus cyclosporine. However, small coronary aneurysms were present at the time of discharge. In a retrospective analysis, no pathogens were detected by multiplex real-time PCR in samples collected at admission, and the serum cytokine profile demonstrated prominent elevation of IL-6 as well as elevation of neopterin, sTNF-RI, and sTNF-RII, which suggested KD. CONCLUSIONS: The patient's entire clinical course, including the severe pneumonia, was caused by KD. As in this case, neonatal KD may exhibit atypical manifestations such as severe pneumonia requiring mechanical ventilation.

Decline in pediatric admission on an isolated island in the COVID-19 pandemic.

BACKGROUND: A decrease in pediatric hospitalizations during the COVID-19 pandemic has been reported worldwide; however, few studies have examined areas with a limited number of COVID-19 cases, where influenced by viral interference by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is minimum. METHODS: We conducted an epidemiological study of pediatric hospitalizations on Sado, an isolated island in Niigata, Japan, that was unique environment with few COVID-19 cases and reliable pediatric admissions monitoring. We compared numbers of monthly hospitalizations and associated diagnoses for the periods April 2016 to March 2020 (pre-pandemic period) and April 2020 to March 2021 (pandemic period). RESULTS: Data were analyzed for 1,144 and 128 patients in the pre-pandemic and pandemic periods, respectively. We observed only three adults and no pediatric COVID-19 cases during the pandemic period. The number of monthly admissions was significantly lower in the pandemic period (median [interquartile ranges (IQR)]: 11.0 [7.0-14.0]) than in the pre-pandemic period (23.0 [20.8-28.3]; P < 0.001). Similar decreases were observed for hospitalizations due to respiratory tract infection (P < 0.01), but not for asthma exacerbation (P = 0.15), and gastrointestinal tract infection (P = 0.33). CONCLUSIONS: Pediatric hospitalizations during the pandemic significantly decreased on an isolated Japanese island where COVID-19 was not endemic and all pediatric admissions were ascertainable. This observation highlights the impact of decreased travel and increased awareness of infection control measures on pediatric hospitalizations due to infectious diseases, not by the SARS-CoV-2 viral interference.

Development of Novel PCR Assays for Improved Detection of Enterovirus D68.

Enterovirus D68 (EV-D68) causes a range of clinical manifestations, including asthma-like illness, severe respiratory disease, and acute flaccid myelitis. EV-D68 has caused worldwide outbreaks since 2014 and is now recognized as a reemerging infection in many countries. EV-D68-specific PCR assays are widely used for the diagnosis of EV-D68 infection; however, assay sensitivity is a concern because of genetic changes in recently circulated EV-D68. To address this, we summarized EV-D68 sequences from previously reported world outbreaks from 2014 through 2020 on GenBank, and found several mutations at the primer and probe binding sites of the existing EV-D68-specific PCR assays. Subsequently, we designed two novel assays corresponding to the recently reported EV-D68 sequences: an EV-D68-specific real-time and seminested PCR. In an analysis of 22 EV-D68 confirmed cases during a recent EV-D68 outbreak in Japan, the new real-time PCR had higher sensitivity than the existing assay (100% versus 45%, P < 0.01) and a lower median CT value (27.8 versus 32.8, P = 0.005). Sensitivity was higher for the new nonnested PCR (91%) than for the existing seminested PCR assay (50%, P < 0.01). The specificity of the new real-time PCR was 100% using samples from non-EV-D68-infected cases (n = 135). In conclusion, our novel assays had higher sensitivity than the existing assay and might lead to more accurate diagnosis of recently circulating EV-D68. To prepare for future EV-D68 outbreaks, EV-D68-specific assays must be continuously monitored and updated.

Innate Immune Responses in Serum and Cerebrospinal Fluid From Neonates and Infants Infected With Parechovirus-A3 or Enteroviruses.

BACKGROUND: Parechovirus (PeV)-A3 and enteroviruses (EV) are the most common viruses causing sepsis and meningoencephalitis in neonates and young infants. Clinical manifestations of PeV-A3 infection are more severe than those of EV infection, and no pleocytosis with a positive polymerase chain reaction (PCR) result for PeV-A3 in cerebrospinal fluid (CSF) are characteristic findings. We hypothesized that innate immune responses to PeV-A3 and EV are distinct in serum and CSF. METHODS: We evaluated 22 cytokines/chemokines in serum and CSF from PeV-A3- or EV-infected patients younger than 4 months in Niigata, Japan, from 2015 through 2018. Infection was diagnosed with real-time PCR followed by sequencing. Febrile neonates and infants with sepsis-like syndrome who had negative bacterial culture and viral PCR for both PeV-A and EV were also included (non-PeV-A/EV patients). RESULTS: Among 192 febrile patients, we evaluated 16 PeV-A3-infected, 15 EV-infected, and 8 non-PeV-A/EV patients. Serum pro-/anti-inflammatory cytokine/chemokine levels were higher in PeV-A3-infected patients than in EV-infected patients (P < .02). Although most cytokine/chemokine were elevated in CSF from EV-infected patients, levels were low or undetectable in PeV-A3-infected and non-PeV-A/EV patients (P < .001). CONCLUSIONS: Distinct cytokine/chemokine patterns in serum and CSF may explain the different clinical manifestations of PeV-A3-infected and EV-infected neonates and young infants.

Parechovirus-A3 encephalitis presenting with focal seizure mimicking herpes simplex virus infection.

BACKGROUND: Febrile neonates and young infants presenting with seizure require immediate evaluation and treatment. Herein we experienced two young infants with parechovirus-A3 (PeV-A3) encephalitis, initially presented with focal seizure suspecting herpes simplex virus (HSV) encephalitis. CASES: We have experienced 2 infantile cases, initially presented with focal seizure. At presentation, HSV encephalitis was strongly suspected and empiric acyclovir therapy was started; however, serum and/or cerebrospinal fluid (CSF) PCR for HSV were negative. Instead, serum and/or CSF PCR for parechovirus-A was positive. PeV-A3 infection was confirmed by genetic sequence analyses. Both cases required multiple anticonvulsant therapy and intensive care for intractable seizure. Diffusion-weighted imaging of brain magnetic resonance imaging (MRI) showed distinct findings; high-intensity lesions in the gray matter of parietal and occipital lobes in Case 1, and bilateral decreased diffusion of the deep white matter and corpus callosum in Case 2. We have followed two cases more than four years; Case 1 developed epilepsy, has been on an anticonvulsant to control her seizure. Case 2 has significant neurodevelopmental delay, unable to stand or communicate with language. CONCLUSIONS: PeV-A3 encephalitis needs to be in differential diagnosis when neonates and young infants present with focal seizure, mimicking HSV encephalitis. Special attention may be necessary in patients with PeV-A3 encephalitis given it could present with intractable seizure with high morbidity in a long-term.

Diagnostic errors in pediatric bacterial osteomyelitis.

BACKGROUND: Clinical evidence for improving diagnostic accuracy in pediatric medicine is still scarce. Septic osteomyelitis is sometimes challenging for physicians to diagnose. The aim of this study was to improve patient care through identifying the incidence and reasons for errors in the diagnosis of bacterial osteomyelitis in pediatric patients. METHODS: We retrospectively identified patients younger than 16 years with acute or chronic osteomyelitis at Tokyo Metropolitan Children's Medical Center between April 2010 and September 2017. We extracted data on patient demographics, clinical course, symptoms, locus of the lesions, and diagnosis at presentation and discharge. The patients were categorized into the misdiagnosis and non-misdiagnosis groups following a review by two pediatricians. Misdiagnosis was defined as a difference between the initial and discharge diagnosis. The factors in the two groups were compared, and the types of error in the misdiagnosis group were examined. RESULTS: In total 71 patients were enrolled. The median age and proportion of boys was 7.6 years (IQR, 1.4-11.2 years) and 66%, respectively. Misdiagnosis occurred in 27 patients (38.0%). Precedent antibiotic use was independently associated with misdiagnosis (P = 0.044). A cognitive error was observed in 88.3% of the misdiagnosis group. The median number of types of error per case was 2.0 (IQR, 2.0-3.0). CONCLUSIONS: The misdiagnosis of septic osteomyelitis in pediatric patients was common and chiefly caused by cognitive errors. Eliminating cognitive errors in diagnosis is highly likely to improve the care of patients with osteomyelitis.

Cefmetazole for extended-spectrum ß-lactamase-producing Enterobacteriaceae in pediatric pyelonephritis.

BACKGROUND: Pyelonephritis caused by extended-spectrum ß-lactamase (ESBL)-producing Enterobacteriaceae is an urgent problem in pediatrics. Although carbapenem is the standard therapy for infections caused by ESBL-producing Enterobacteriaceae, some cephamycins, including cefmetazole, are stable against hydrolysis by ESBL. There are few reports, however, on the use of cefmetazole in children. The aim of this study was to evaluate the therapeutic effect of cefmetazole in pediatric pyelonephritis caused by ESBL-producing Enterobacteriaceae. METHODS: Children with pyelonephritis caused by ESBL-producing Enterobacteriaceae were enrolled between April 2010 and November 2016 at Tokyo Metropolitan Children's Medical Center. Presence of ESBL was tested for using the disk diffusion method. Medical records were reviewed for a past history of bacterial infection. The outcomes were clinical cure rate at 4 weeks and the duration of therapy in the cefmetazole and non-cefmetazole groups. RESULTS: Fifty-five patients met the criteria for pyelonephritis caused by ESBL-producing Enterobacteriaceae. The most common causative organisms were Escherichia coli (n = 51; 92.7%), Klebsiella pneumoniae (n = 3; 5.5%), and K. oxytoca (n = 1; 1.8%). Thirty-six and 19 patients were treated with cefmetazole and with other antibiotics as definitive therapy, respectively. There was no difference in the clinical cure rate (86.1% vs 89.5%; P = 0.72) or duration of therapy (median, 7.0 vs 7.0 days; P = 0.73) between the cefmetazole and non-cefmetazole groups. CONCLUSIONS: Cefmetazole was not inferior to the other antibiotics in the treatment of pyelonephritis caused by ESBL-producing Enterobacteriaceae in children. Cefmetazole is a valuable therapeutic alternative to carbapenems for treating pyelonephritis caused by ESBL-producing Enterobacteriaceae.

Sibling visits and viral infection in the neonatal intensive care unit.

BACKGROUND: Sibling visits to the neonatal intensive care unit (NICU) are a part of family-centered care, which is now being increasingly endorsed as a positive development in patient care. Sibling visits, however, pose a risk of viral infection, and hence many NICU in Japan impose strict limits on the practice. The aim of this study was therefore to assess whether sibling visits to the NICU are related to an increase in the nosocomial viral infection rate. METHODS: This retrospective study was conducted between April 2012 and March 2017 at Tokyo Metropolitan Children's Medical Center in Japan. Sibling visits were implemented after screening for symptoms of viral illness. Symptomatic patients in the NICU were tested for common viruses on rapid antigen test and polymerase chain reaction. The number of sibling visits and the rate of nosocomial viral infections were examined on Spearman's correlation test. RESULTS: The total number of sibling visits and rate of nosocomial viral infection in the NICU was 102 and 0.068 per 1,000 patient-days during the study period, respectively. The number of enterovirus, respiratory syncytial virus, human metapneumovirus, influenza virus A, and Herpes simplex virus infections was 3, 2, 1, 1, 1, and 1, respectively. No infections were identified after sibling visits. The number of sibling visits and the rate of nosocomial viral infections were not correlated (correlation coefficient, -0.1; P = 0.873). CONCLUSION: Sibling visits to the NICU did not result in an increase in the nosocomial viral infection rate.

Human parechovirus type 3 infection: An emerging infection in neonates and young infants.

Human parechoviruses (HPeVs) are RNA viruses that have characteristics similar to those of enteroviruses and usually cause mild respiratory or gastrointestinal symptoms. Human parechovirus type 3 (HPeV3), first reported in 2004, is exceptional because it can provoke sepsis and meningoencephalitis leading to neurological sequelae, and even death, in neonates and young infants. Pediatricians and researchers are increasingly aware that HPeV3 is responsible for serious disease in neonates and young infants. Retrospective studies and several reports of epidemics of HPeV3 infection have provided data on epidemiology, clinical symptoms and signs, laboratory findings, and outcomes. However, the pathogenesis of HPeV3 infection remains unclear, which explains the lack of specific antiviral therapy and effective prevention measures. Maternal antibodies are important in protection against severe HPeV3-related disease, and this may be a clue regarding its pathogenesis. HPeV3 epidemics are likely to continue, and because the clinical manifestations of HPeV3 are severe, determining the pathogenesis of HPeV3 infection and establishing specific antiviral therapies are important goals for future research.

Human parechovirus type 3 infection: Cause of apnea in infants born prematurely.

Four infants born prematurely presented with multiple apnea episodes caused by human parechovirus type 3 (HPeV3) infection. All patients required oxygen supplementation, and one patient required mechanical ventilation. HPeV3 infection might be included in the differential diagnosis of apnea in neonates and young infants, especially those born prematurely.

Role of Maternal Antibodies in Infants with Severe Diseases Related to Human Parechovirus Type 3.

Human parechovirus type 3 (HPeV3) is an emerging pathogen that causes sepsis and meningoencephalitis in young infants. To test the hypothesis that maternal antibodies can protect this population, we measured neutralizing antibody titers (NATs) to HPeV3 and other genotypes (HPeV1 and HPeV6) in 175 cord blood samples in Japan. The seropositivity rate (≥1:32) for HPeV3 was 61%, similar to that for the other genotypes, but decreased significantly as maternal age increased (p<0.001). Furthermore, during the 2014 HPeV3 epidemic, prospective measurement of NATs to HPeV3 in 45 patients with severe diseases caused by HPeV3 infection showed low NATs (≤1:16) at onset and persistently high NATs (≥1:512) until age 6 months. All intravenous immunoglobulin samples tested elicited high NATs to HPeV3. Our findings indicate that maternal antibodies to HPeV3 may help protect young infants from severe diseases related to HPeV3 and that antibody supplementation may benefit these patients.