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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. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Background Enterovirus D68 (EV-D68) is a re-emerging, high-morbidity pathogen that causes severe respiratory infection and acute flaccid myelitis in children. EV-D68 is phylogenetically divided into 4 major groups—clade A through D—and each clade has subclades defined by genetic variant. Outbreaks of all strains have been reported in more than 10 countries. However, no study has compared the viral genomic characteristics of EV-D68 in contemporaneous outbreaks in different countries. Methods An outbreak of EV-D68 in children younger than 15 years occurred in Niigata, Japan, from October through November 2018. The patients were admitted to hospital with respiratory distress and wheezing episodes. RNA extracted from nasopharyngeal samples was tested by EV-D68-specific PCR. Clade was determined by semi-nested PCR analysis of the VP1 region, and the phylogenetic tree of the VP1 sequence was constructed. To clarify viral genomic characteristics, we compared the clade to those of outbreaks occurring during 2014–2018 in other countries. Results EV-D68 testing of 47 patients yielded positive results for 22 (47%) (median age 4.6 years; IQR, 2.9–6.7), and 15 (69%) had a past medical history of allergic disease. No patient developed acute flaccid myelitis. The VP1 sequences from all isolates belonged to clade B3, the same clade detected during the 2015 outbreak in Japan (figure). Interestingly, EV-D68 outbreaks were reported in France and Italy (clades D1/B3 and D1, respectively) during this period. Although the EV-D68 clades responsible for outbreaks in 2014 differed by region, clade B3 is now the dominant clade and has caused concurrent EV-D68 outbreaks in children since 2015 (table). Conclusion In 2018, an EV-D68 clade B3 outbreak occurred among children in Niigata, Japan. This clade is now dominant and periodically circulates around the world. Because EV-D68 can cause simultaneous outbreaks worldwide and is associated with high morbidity, detailed real-time epidemiological surveillance of EV-D68 is warranted. Disclosures All authors: No reported disclosures. [ABSTRACT FROM AUTHOR]

Background Parechovirus-A3 (PeV-A3) causes severe disease, including sepsis and meningoencephalitis in young infants. The first case of PeV-A3 was reported in Japan in 1999 and, although epidemics have been reported every 2 to 3 years in more than 20 countries, no major epidemic has occurred in Japan since 2014. Methods This prospective study included febrile infants (<4 months of age) admitted at Niigata University and its affiliated hospitals, which serve about 2.5 million people, during the period from 2014 to 2018. Neonates and infants younger than 4 months presenting with fever and suspected of having viral sepsis underwent serum and/or cerebrospinal fluid (CSF) testing by real-time PCR for parechovirus-A (PeV-A) and enteroviruses (EVs), and for herpes simplex viruses, if suspected. Bacterial infection was excluded on the basis of the results of bacterial culture of blood, urine, and/or CSF. PeV-A genotype was identified by examining the viral protein 1 (VP1) sequence, and the phylogenetic tree of the VP1 sequence was constructed. Results Of the 277 patients evaluated, 135 (49%) were positive for PeV-A (n = 74, 27%) or EVs (n = 61, 22%). Among PeV-A patients, most had PeV-A3 (n = 69; 93%), followed by PeV-A4 (n = 4; 5%). There was a PeV-A3 epidemic in 2014 (n = 43); however, no cases were reported in 2015. In 2016–2018, small numbers of PeV-A3 cases were reported: 10 in 2016, 7 in 2017, and 9 in 2018. In contrast, EV cases were reported throughout this period: 8 in 2014, 22 in 2015, 10 in 2016, 5 in 2017, and 16 in 2018. When data were analyzed by season, the PeV-A3 detection rate in summer (June-August) was 93% (40/43) in 2014 and 65% (17/26) during 2016–2018, indicating an increase in the number of PeV-A3 cases in seasons other than summer. Phylogenetic analysis showed that PeV-A3 strains during 2016–2018 were part of a cluster of epidemics in 2011 and differed from those in 2014. Conclusion After the major PeV-A3 epidemics in 2014, we observed changes in the PeV-A3 epidemic profile, namely, a small, but constant, number of cases every year in Niigata, Japan. A future study should investigate if this trend has continued in Japan and other countries and identify the causes of this change in epidemic profile. Disclosures All authors: No reported disclosures. [ABSTRACT FROM AUTHOR]

• HPeV-3 and EV are commonly detected in febrile neonates and infants. • Serum and CSF were PCR-positive for 100 % and 88 % of HPeV-3-infected patients, respectively. • Serum and CSF were PCR-positive for 83 % and 71 % of EV-infected patients, respectively. • In EV-infected patients, detection rates for serum and CSF varied by EV genotype. • Collection of both serum and CSF samples improves viral diagnosis in this population. Human parechovirus 3 (HPeV-3) and enteroviruses (EV) are commonly detected viruses in febrile neonates and young infants and are usually diagnosed by PCR. However, in this population, data on detection rates for samples from different anatomical sites are limited. To determine PCR detection rates for HPeV-3 and EVs in serum and cerebrospinal fluid (CSF) samples from febrile neonates and young infants. This prospective study identified viruses in serum and CSF samples collected from febrile neonates and young infants (age <4 months) in Niigata, Japan, during 2014−2018. HPeV-3 or EV infection was defined as a positive quantitative real-time PCR result for the virus in serum or CSF. Genotypes were identified by sequence analyses of the viral protein 1 region. Among 216 patients, we identified 56 HPeV-3-infected (26 %) and 48 EV-infected patients (22 %). All (56/56; 100 %) HPeV-3-infected patients had a positive PCR result for serum, and 49/56 (88 %) had a positive result for CSF. In EV-infected patients, 40/48 (83 %) were positive for serum, and 34/48 (71 %) were positive for CSF, and 22/48 (46 %) were positive for serum (n = 14) or CSF (n = 8). If only a CSF sample had been obtained, 7 (12 %) HPeV-3 infections and 14 (29 %) EV infections would have been undiagnosed. Detection rates in serum and CSF differed by genotype in EV-infected patients. Viral RNA detection rates differed between serum and CSF in HPeV-3- and EV-infected neonates/infants. Combined evaluation of serum and CSF samples is important for accurate viral diagnosis in this population. [ABSTRACT FROM AUTHOR]

• PeV-A3 and EVs are the most common viruses causing severe diseases in young infants. • PeV-A3 patients had some severer symptoms and signs compared to EVs patients. • PeV-A3 patients tended to have a lower WBC count in blood and no CSF pleocytosis. • The scoring system with these findings were successful to distinguish 2 infections. Parechovirus-A3 (PeV-A3) and the enteroviruses (EVs) are the most common viral pathogens responsible for sepsis and meningoencephalitis in neonates and young infants; however, differences in the clinical presentations of two infections are not well described. To describe the clinical presentations of PeV-A3- and EVs-related diseases and develop a novel scoring system to differentiate two diseases. This prospective study used real-time PCR and genetic sequencing to evaluate viral etiologies of febrile neonates and infants <4 months with suspected sepsis or meningoencephalitis in Niigata area, Japan, in 2014-2016. The clinical manifestations of PeV-A3- and EVs-infected patients were compared, and a novel scoring system was developed after identifying the most distinguishable clinical findings, followed by the external cohort validation. In 210 patients evaluated, we identified 56 PeV-A3-infected (27%) and 43 EVs-infected (20%) patients. The following clinical manifestations were significant in PeV-A3-infected patients, as compared with EVs-infected patients; a higher body temperature (38.9°C vs. 38.5°C, P <.01) and heart rate (181/min vs. 168/min, P =.01), cold extremities (72% vs. 34%, P <.01) and skin mottling (65% vs. 23%, P <.01), lower white blood cell count (5,200/μL vs. 8,900/μL, P <.01) and incidence of cerebrospinal fluid (CSF) pleocytosis (2% vs. 63%, P <.01). Using some of these significant findings, the scoring system successfully distinguished the diseases (accuracy: 86% and 83% for the derivative and external validation cohorts, respectively). We found significant clinical manifestations in PeV-A3-infected patients compared to EVs-infected patients. The scoring system may be helpful to distinguish two infections, especially at onset of outbreak. [ABSTRACT FROM AUTHOR]

Background Parechovirus-A3 (PeV-A3) and enteroviruses (EVs) are the most common viral causes of neonatal and infantile sepsis. We previously reported that the clinical manifestations of PeV-A3 infection—e.g. high body temperature, tachycardia, and poor peripheral circulation, but not cerebrospinal fluid (CSF) pleocytosis—tend be more severe than those of EV infection. We tested the hypothesis that innate immune responses to PeV-A3 and EVs are distinct. Methods Using serum and CSF samples, we investigated immune responses of febrile neonates and infants <4 months in Niigata, Japan, from 2015 through 2018. PeV-A and EV infections were diagnosed with real-time PCR. PeV-A3 infection was diagnosed by sequence analysis of the VP1 region. The control was clinically well patients without serum and CSF findings suggestive of bacterial or viral etiology. The Milliplex MAP human cytokine/chemokine magnetic bead panel (Merck Millipore, Germany) was used to analyze 22 cytokines/chemokines related to innate immunity in serum and CSF. Results We evaluated 14 PeV-A3-infected and 15 EV-infected patients and 8 controls. Serum levels of proinflammatory cytokines/chemokines (fractalkine, interferon-α2, interleukin [IL]-1 receptor α, IL-6, IL-8, and IL-15) were significantly higher in PeV-A3-infected patients than in EV-infected patients (P < 0.005). Serum cytokine/chemokine profiles of EV-infected patients did not differ from those of controls. However, while most pro- and anti-inflammatory cytokines/chemokines in CSF were elevated in EV-infected patients, levels were low or undetectable in PeV-A3-infected patients and controls (P < 0.005). Conclusion PeV-A3-infected patients had high serum levels of proinflammatory cytokines/chemokines, which may explain why clinical manifestations were more severe in this patient group than in EV-infected patients. Conversely, the limited or nonexistent innate immune response in CSF from PeV-A3-infected patients might explain the absence of CSF pleocytosis. These findings improve our understanding of the differing pathophysiological characteristics of PeV-A3 and EV infection in neonates and young infants. Disclosures All Authors: No reported Disclosures. [ABSTRACT FROM AUTHOR]

Background Parechovirus-A3 (PeV-A3) is an emerging pathogen causing sepsis and meningoencephalitis in neonates and young infants. We previously reported that maternal antibodies against PeV-A3 are important to protect neonates and young infants from the infection. Recent studies showed that (1) breastfeeding had a protective effect against enterovirus, which is closely-related virus to PeV-A, and (2) human breast milk (HBM) neutralized enterovirus in vitro. Currently, no report is available related to the antiviral effect of HBM against PeV-A3. Methods HBM (colostrum, 3–5 days after childbirth; mature milk, 1 month after childbirth) and serum (within ± 1 week of child's birthday) samples were obtained from mothers at obstetrics clinic in Niigata, Japan. Neutralizing antibody titers (NATs) against PeV-A3 were measured using the Vero cells. Results The anti-PeV-A3 NATs of colostrum (n = 32) ranged from 1:8 to 1:2048, those ≥1:32 was 59% (19/32). Whereas, the anti-PeV-A3 NATs of mature milk ranged from 1:8 to 1:96. and those ≥1:32 was 20% (2/20) (P < 0.001). The median NATs anti-PeV-A3 was higher in colostrum (1:32) compared with mature milk (1:8) (P < 0.001). There was a strong positive correlation between the NATs of colostrum and serum (r = 0.604, P < 0.001, Figure). Conclusion This study showed that HBM had high NATs against PeV-A3, which was correlated with serum NATs. Further studies are necessary to investigate which components of HBM has antiviral effects against PeV-A3. Disclosures All authors: No reported disclosures. [ABSTRACT FROM AUTHOR]