Human parechovirus (HPeV) is a small non-enveloped RNA virus that belongs to the family Picornaviridae. It was first isolated in 1956 and the first two viruses were classified as echoviruses 22 and 23 of the genus Enterovirus (EV). In 1999 they were reclassified into the genus Parechovirus, as HPeV-1 and HPeV-2 respectively. Currently, 19 genotypes have been identified, being HPeV-3 particularly neurotropic and associated with more severe disease¹⁰.

Infection is prevalent among young children. It is often asymptomatic or associated with mild disease of the gas trointestinal and respiratory tracts. However, HPeV can also cause severe diseases such as sepsis, encephalitis, and meningitisith long term neurological sequelae in young infants, including cerebral palsy, central visual impairment, and gross motor impairmentTherefore, it represents a diagnostic and therapeutic challenge for the treating physi- cian.

HPeV detection is not routinely performed in most clinical microbiology laboratories and, therefore, its real preva- lence is unknown. Recent studies have shown that HPeV-3 might be one of the main agents involved in neonatal neu- rological infections worldwide

We report three cases that presented with different clin- ical features in our hospital after the implementation of a multiplex PCR meningitis/encephalitis panel (ME) (Filmar- ray) between December 2018 and December 2019. The panel includes the detection of 7 viruses (HPeV, EV, herpes simplex virus 1 and 2, human herpesvirus 6, varicella-zoster virus), 6 bacteria (Escherichia coli K1, Haemophilus influenzae, Listeria monocytogenes, Neisseria meningitidis, Streptococ- cus agalactiae, Streptococcus pneumoniae) and one yeast (Cryptococcus neoformans/gattii).

A 6-day-old boy was brought to the Department of Pedi- atrics for a neonatal jaundice follow-up examination. His mother reported decreased oral intake and a febrile episode of 37.9 °C without gastrointestinal or respiratory symptoms. He was cesarean-born and his mother had a Streptococ- cus group B (SGB)-positive culture during pregnancy. His mother refused to hospitalize him and returned the follow- ing day reporting fever, irritability, and decreased feeding. He also presented with mild aqueous diarrhea that was self-limiting. He was admitted to the neonatal ICU. Labo- ratory tests included lumbar puncture, urine, stool, and a set of blood cultures. His white blood cell count (WBCc) was 8.7 x10³WBC/mm³ and his C-reactive protein (CRP) level was 1.6mg/l. He was administered intravenous ampicillin and gentamicin. The cerebrospinal fluid (CFS) was clear and colorless, had no pleocytosis, only mild proteinorrachy (87 mg/dl). Bacterial cultures of blood, urine, and CFS were negative and his chest X-ray was normal. The ME panel was positive for HPeV and rotavirus antigen was detectable in the stool. On day 2 of admission, he developed left tonic-clonic movements and episodes of bradycardia and desaturation. Phenobarbital was administered (15mg/kg). Brain ultra- sound showed subcortical white matter hypoechogenicity of both cerebral hemispheres, corpus callosum, and both tem poral lobes. Brain magnetic resonance imaging (MRI) showed acute leukoencephalitis (Fig. 1). He also presented an ery- thematous palmar-plantar rash (Fig. 2). Levetiracepam was added to his treatment scheme as he manifested new desat- uration episodes and hypertonia. He gradually improved and was discharged on day 15. Due to the severity of the case, HPeV was identified by real-time RT-PCR as HPeV-3. He is currently followed-up in Child Neurology, continues on an anticonvulsant treatment, and receives early stimulation.

Figure 1: Case 1 - axial magnetic resonance imaging. Diffusion sequence (a) and apparent diffusion coefficient (ADC) map (b) where extensive restrictive areas of diffuse and symmetric distribution are observed, compromising the supratentorial bihemispheric white matter, dorso-lateral nuclei of both thalamus and corpus callosum. T2 * Sequence (c) showing the bleeding area (red arrow). 

Figure 2: Case 1 - erythematous plantar rash that presented in our hospital between December 2018 and December 2019. 

A 16-day-old girl was admitted to the emergency depart- ment for a history of decreased oral intake, irritability, and fever of 38 °C without gastrointestinal or respiratory symptoms. She was born full-term by vaginal delivery without complications and her mother had an SGB-positive culture during pregnancy. On physical examination, no rash was observed. Laboratory tests included lumbar punc- ture, urine, and a set of blood cultures. Her WBCc was 6.2 x 10³ WBC/mm³ and her CRP level was 2.0mg/l. She was administered intravenous gentamicin and cefotaxime. Her CFS was clear and colorless and without pleocytosis. Bacte rial cultures of blood, urine, and CFS were negative. The ME panel was positive for HPeV. Her chest X-ray was normal and a nasopharyngeal swab was negative for respiratory viruses. She was admitted for 3 days, and after 24 h with no fever and a favorable evolution, she was discharged.

A 4-month-old boy, born full-term by vaginal delivery, was admitted to the emergency department after 4 h of per- sistent crying and two febrile episodes of 38 °C in the last 24 h. He had normal weight-for-age. On physical examina- tion, he was irritable with abdominal tenderness and no rash was observed. His brother was having an upper respiratory tract infection. His WBCc was 6.2 x 10³ WBC/mm³ and his CRP level was 5.2mg/l. Abdominal ultrasound only showed meteorism. A lumbar puncture was performed, and CFS was clear and colorless, without pleocytosis. The ME panel was positive for HPeV. Bacterial cultures of blood, urine, and CFS were negative. A cerebral ultrasound was performed and was normal. He had intermittent sub-febrile episodes and was discharged on day 4 of admission, after 30 h with no fever.

HPeV is one of the picornaviruses that has been associ- ated with fever of unknown origin, gastroenteritis, clinical sepsis, meningitis, or encephalitis in very young infants. We report three different cases of HPeV CNS that presented in our hospital between December 2018 and December 2019.

HPeV infections have been reported worldwide, and pre- vious studies have shown a prevalence of HPeV in CSF samples ranging from 0 to 17%Gutierrez et al. reported HPeV as the etiologic agent of meningitis and sepsis-like ill- ness in infants with a detection rate of 18% in South America, which is higher than any other series worldwideHowever, the prevalence rates differ depending on the design of the study, sampling sites, and demographic characteristics. As HPeV infection is not a notifiable disease in Argentina and the test is not widely accessible, epidemiological informa- tion is not available. For the same age group in the studied period, the incidence of HPeV and EV infections was simi lar, as there were three viral encephalitis cases due to EV. We, therefore, conclude that further studies are needed to estimate the burden of HPeV infections.

The three cases were recorded during the spring, as it has been previously describedHowever, most HPeV infections tend to occur in late summer and autumn monthsThe age distribution of patients with HPeV infection is characterized for presenting in children under three months of age, and ainly in neonatesIn this cohort, the three patients were less than 4 months of age.

Transmission of HPeV occurs through the fecal-oral route and possibly infective respiratory dropletsThere is scant data regarding respiratory transmission, however, it is sup- posed to be the route in children with CNS symptomsThe risk of transmission is probably highest during acute symp- tomatic illness. Confirmed cases should be under standard, contact, and respiratory precautions to prevent transmission to other young infants during admission

All cases presented with fever and irritability as common clinical features. However, the severity of case 1 was quite different, reflecting the wide range of clinical features of this viral infection. Although most mild HPeV infections have no specific symptoms, severe infections have some clinical features that may suggest the etiology. Clinical syndromes caused by HPeV vary by genotype, and HPeV-3 illness can be severe with up to 20-50% of admitted patients present ing with sepsis-like syndromeThe combination of fever, rash, and severe irritability with characteristics of sepsis in neonates should lead clinicians to consider HPeV infection as a differential diagnosisLong term follow-up is strongly advised, as neonates are considered a vulnerable popula- tion at risk for neurodevelopmental delay and neurological sequelae

The absence of CSF pleocytosis and normal protein and glucose levels in CSF are commonly reported in most cases of HPeV CNS infections, as observed in these three casesHPeV should be considered for testing in CFS samples of neonates as it can lead to severe encephalitis associated with white matter injury in the neonatal periodSyndromic molec ular testing allowed us to establish the specific cause of encephalitis at around 3 h since sample collection. A shorter time to result could lead to shorter hospital stays and a decrease in the use of antimicrobial therapy

Molecular diagnostic techniques could help improve patient care and understand the real prevalence of this infection in Argentina.

Conflict of interest

The authors declare that they have no conflicts of interest.B. Fox, V. Sabio Paz, M.A. Incardona et al.

Acknowledgments

We thank María Cecilia Freire, Servicio de Neurovirosis, INEI- ANLIS Dr. Carlos G. Malbran, for genotyping HPeV-3 from case