N6-methyladenosine modification of a parvovirus-encoded small noncoding RNA facilitates viral DNA replication through recruiting Y-family DNA polymerases.

Human bocavirus 1 (HBoV1) is a human parvovirus that causes lower respiratory tract infections in young children. It contains a single-stranded (ss) DNA genome of ~5.5 kb that encodes a small noncoding RNA of 140 nucleotides known as bocavirus-encoded small RNA (BocaSR), in addition to viral proteins. Here, we determined the secondary structure of BocaSR in vivo by using DMS-MaPseq. Our findings reveal that BocaSR undergoes N6-methyladenosine (m6A) modification at multiple sites, which is critical for viral DNA replication in both dividing HEK293 cells and nondividing cells of the human airway epithelium. Mechanistically, we found that m6A-modified BocaSR serves as a mediator for recruiting Y-family DNA repair DNA polymerase (Pol) η and Pol κ likely through a direct interaction between BocaSR and the viral DNA replication origin at the right terminus of the viral genome. Thus, this report represents direct involvement of a viral small noncoding RNA in viral DNA replication through m6A modification.

Sequencing and characterization of human bocavirus genomes from patients diagnosed in Southern France between 2017 and 2022.

The diversity and evolution of the genomes of human bocavirus (HBoV), which causes respiratory diseases, have been scarcely studied. Here, we aimed to obtain and characterize HBoV genomes from patients's nasopharyngeal samples collected between 2017 and 2022 period (5 years and 7 months). Next-generation sequencing (NGS) used Illumina technology after having implemented using GEMI an in-house multiplex PCR amplification strategy. Genomes were assembled and analyzed with CLC Genomics, Mafft, BioEdit, MeV, Nextclade, MEGA, and iTol. A total of 213 genomes were obtained. Phylogeny classified them all as of Bocavirus 1 (HBoV1) species. Five HBoV1 genotypic clusters determined by hierarchical clustering analysis of 27 variable genome positions were scattered over the study period although with differences in yearly prevalence. A total of 167 amino acid substitutions were detected. Besides, coinfection was observed for 52% of the samples, rhinoviruses then adenoviruses (HAdVs) being the most common viruses. Principal component analysis showed that HBoV1 genotypic cluster α tended to be correlated with HAdV co-infection. Subsequent HAdV typing for HBoV1-positive samples and negative controls demonstrated that HAdVC species predominated but HAdVB was that significantly HBoV1-associated. Overall, we described here the first HBoV1 genomes sequenced for France. HBoV1 and HAdVB association deserves further investigation.

T84 Monolayer Cell Cultures Support Productive HBoV and HSV-1 Replication and Enable In Vitro Co-Infection Studies.

Based on several clinical observations it was hypothesized that herpesviruses may influence the replication of human bocaviruses, the second known parvoviruses that have been confirmed as human pathogens. While several cell lines support the growth of HSV-1, HBoV-1 was exclusively cultivated on air-liquid interface cultures, the latter being a rather complicated, slow, and low throughput system. One of the cell lines are T84 cells, which are derived from the lung metastasis of a colorectal tumor. In this study, we provide evidence that T84 also supports HBoV replication when cultivated as monolayers, while simultaneously being permissive for HSV-1. The cell culture model thus would enable co-infection studies of both viruses and is worth being optimized for high throughput studies with HBoV-1. Additionally, the study provides evidence for a supporting effect of HSV-1 on the replication and packaging of HBoV-1 progeny DNA into DNase-resistant viral particles.

Presence and Significance of Multiple Respiratory Viral Infections in Children Admitted to a Tertiary Pediatric Hospital in Italy.

Viral co-infections are frequently observed among children, but whether specific viral interactions enhance or diminish the severity of respiratory disease is still controversial. This study aimed to investigate the type of viral mono- and co-infections by also evaluating viral correlations in 3525 respiratory samples from 3525 pediatric in/outpatients screened by the Allplex Respiratory Panel Assays and with a Severe Acute Respiratory Syndrome-COronaVirus 2 (SARS-CoV-2) test available. Overall, viral co-infections were detected in 37.8% of patients and were more frequently observed in specimens from children with lower respiratory tract infections compared to those with upper respiratory tract infections (47.1% vs. 36.0%, p = 0.003). SARS-CoV-2 and influenza A were more commonly detected in mono-infections, whereas human bocavirus showed the highest co-infection rate (87.8% in co-infection). After analyzing viral pairings using Spearman's correlation test, it was noted that SARS-CoV-2 was negatively associated with all other respiratory viruses, whereas a markedly significant positive correlation (p < 0.001) was observed for five viral pairings (involving adenovirus/human bocavirus/human enterovirus/metapneumoviruses/rhinovirus). The correlation between co-infection and clinical outcome may be linked to the type of virus(es) involved in the co-infection rather than simple co-presence. Further studies dedicated to this important point are needed, since it has obvious implications from a diagnostic and clinical point of view.

Human Bocavirus 1 NP1 acts as an ssDNA-binding protein to help AAV2 DNA replication and cooperates with RPA to regulate AAV2 capsid expression.

Adeno-associated virus (AAV) requires co-infection with helper virus for efficient replication. We previously reported that Human Bocavirus 1 (HBoV1) genes, including NP1, NS2, and BocaSR, were critical for AAV2 replication. Here, we first demonstrate the essential roles of the NP1 protein in AAV2 DNA replication and protein expression. We show that NP1 binds to single-strand DNA (ssDNA) at least 30 nucleotides (nt) in length in a sequence-independent manner. Furthermore, NP1 colocalized with the BrdU-labeled AAV2 DNA replication center, and the loss of the ssDNA-binding ability of NP1 by site-directed mutation completely abolished AAV2 DNA replication. We used affinity-tagged NP1 protein to identify host cellular proteins associated with NP1 in cells cotransfected with the HBoV1 helper genes and AAV2 duplex genome. Of the identified proteins, we demonstrate that NP1 directly binds to the DBD-F domain of the RPA70 subunit with a high affinity through the residues 101-121. By reconstituting the heterotrimer protein RPA in vitro using gel filtration, we demonstrate that NP1 physically associates with RPA to form a heterologous complex characterized by typical fast-on/fast-off kinetics. Following a dominant-negative strategy, we found that NP1-RPA complex mainly plays a role in expressing AAV2 capsid protein by enhancing the transcriptional activity of the p40 promoter. Our study revealed a novel mechanism by which HBoV1 NP1 protein supports AAV2 DNA replication and capsid protein expression through its ssDNA-binding ability and direct interaction with RPA, respectively.IMPORTANCERecombinant adeno-associated virus (rAAV) vectors have been extensively used in clinical gene therapy strategies. However, a limitation of these gene therapy strategies is the efficient production of the required vectors, as AAV alone is replication-deficient in the host cells. HBoV1 provides the simplest AAV2 helper genes consisting of NP1, NS2, and BocaSR. An important question regarding the helper function of HBoV1 is whether it provides any direct function that supports AAV2 DNA replication and protein expression. Also of interest is how HBoV1 interplays with potential host factors to constitute a permissive environment for AAV2 replication. Our studies revealed that the multifunctional protein NP1 plays important roles in AAV2 DNA replication via its sequence-independent ssDNA-binding ability and in regulating AAV2 capsid protein expression by physically interacting with host protein RPA. Our findings present theoretical guidance for the future application of the HBoV1 helper genes in the rAAV vector production.

[Construction and characterization of an infectious clone for human bocaparvovirus HBoV1].

Human bocaparvovirus 1 (HBoV1) is one of the two parvoviruses that infect humans and cause diseases. Infection with HBoV1 in infants and young children aged 2-5 years can lead to mild or severe acute respiratory diseases, with the most severe cases posing a life-threatening risk. Similar to other parvoviruses, the HBoV1 DNA genome consists of two terminal reverse repeats (ITRs) at its ends, which are necessary for viral genome replication. However, up to now, it has remained a technical challenge to clone the entire ITRs through PCR amplification. In this study, we successfully constructed a full-length infectious clone of HBoV1, termed as pSKHBoV1, by synthesizing and cloning the terminal ITRs in a stepwise manner. After transfecting HEK293 cells with the infectious clone pSKHBoV1, we were able to reconstitute the viral replication cycle. This included the expression of key non-structural proteins, post-transcriptional modification and processing of viral RNA, viral genome replication, and potentially the production of progeny virions containing the defined DNA genome. The successful construction of the infectious clone pSKHBoV1 lays the foundation for future studies on HBoV1 replication and propagation, virus-host interaction, and the development of viral vaccines.

Molecular characterization of human bocavirus in municipal wastewaters using amplicon target sequencing.

Human bocavirus (HBoV) is an emerging health concern worldwide, associated with range of clinical manifestations, including gastroenteritis and respiratory infections. Therefore, it is crucial to comprehend and minimize their prevalence in different systems. In this study, we conducted regular sampling throughout the year in two different sizes and work processes of wastewater treatment plants (WWTPs) in Tianjin, China. Our objective was to investigate the occurrence, prevalence, and endurance of HBoV in wastewater, while also evaluating the efficacy of amplicon target sequencing in directly detecting HBoV in wastewater. At two WWTPs, HBoV2 (45.51 %-45.67 %) and HBoV3 (38.30 %-40.25 %) were the most common genotypes identified, and the mean concentration range of HBoV was 2.54-7.40 log10 equivalent copies/l as determined by multiplex real-time quantitative PCR assay. A positive rate of HBoV was found in 96.6 % (29/30) samples of A-WWTP, and 96.6 % (26/27) samples of B-WWTP. The phylogenetic analysis indicated that the nucleotide similarity between the HBoV DNA sequences to the reference HBoV sequences published globally ranged from 90.14 %-100 %. A significant variation in the read abundance of HBoV2 and HBoV3 in two wastewater treatment plants (p < 0.05) was detected, specifically in the Winter and Summer seasons. The findings revealed a strong correlation between the genotypes detected in wastewater and the clinical data across various regions in China. In addition, it is worth mentioning that HBoV4 was exclusively detected in wastewater and not found in the clinical samples from patients. This study highlights the high prevalence of human bocavirus in municipal wastewater. This finding illustrates that amplicon target sequencing can amplify a wide variety of viruses, enabling the identification of newly discovered viruses.

Genotypic diversity and long-term impact of human bocavirus on diarrheal disease: Insights from historical fecal samples in Brazil.

This study aimed to investigate the frequency and genotypic diversity of human bocavirus (HBoV) in historical fecal samples collected before 2005 in Brazil and understand its natural history in patients with diarrhea. Between 1998 and 2005, 3347 samples were tested for HBoV by RT-PCR, with a detection rate of 5.8% (195/3347). Coinfection with norovirus (NoV) and human adenovirus (HAdV) was found in 34.9% (68/195), indicating HBoV's potential role as a causative agent of diarrheal disease. The detection rate varied over the years (p < 0.05), suggesting natural oscillatory fluctuations. HBoV was more prevalent in fall and winter, with higher positivity in children ≤5 years (p < 0.05), reinforcing that HBoV is an important pathogen in childhood diarrhea. Genotyping (32.8%; 64/195) revealed the circulation of HBoV-1 (79.7%, 51/64), HBoV-3 (12.5%, 8/64), HBoV-2 (6.2%, 4/64), and the rare HBoV-4 (1.6%, 1/64). Difference in HBoV-1 and HBoV-2/-3 mono-infections prevalence (p < 0.05), suggests a potential role of HBoV-1 in the pathogenicity of diarrheal disease. The study highlights HBoV's lasting impact on viral gastroenteritis in Brazil and emphasizes its genotypic diversity. Recommending screening for HBoV in public health laboratories is crucial for understanding its role in gastrointestinal diseases. The data also contribute to understanding the molecular characterization of enteric viruses in historical fecal samples.

Bocaparvovirus in infants hospitalized in a neonatal intensive care unit in Midwest Brazil during the COVID-19 pandemic (2021-2022).

Human bocaparvoviruses (HBoVs) belong to the Parvoviridae family, being currently classified into four species (HBoV1-4). These viruses have been found in association with respiratory and gastroenteric symptoms, as well as in asymptomatic individuals. This study aimed to investigate the occurrence of HBoVs in infants under 5 months old admitted to a Neonatal Intensive Care Unit (NICU) during the COVID-19 pandemic (between March 2021 and March 2022). Clinical samples (nasopharyngeal swab, serum, stool, and urine) were screened by qPCR TaqMan. The HBoV was detected in samples of 31.6% (12/38) of participants. The most frequent alteration among the HBoV-positive neonates was the chest X-ray with interstitial infiltrate, followed by tachycardia and vomiting. Viral DNA was detected in more than one type of clinical sample in three of the participants in association with respiratory symptoms. Two participants had positive stool samples with or without enteric symptoms. HBoV intermittent and continuous positivity patterns were observed. The present study stands out for the prospective evaluation of positivity for HBoV in different types of clinical samples from a population of hospitalized infants. Our data supports circulation of HBoV in nosocomial environment during the COVID-19 pandemic.

Human Bocavirus infection in childhood acute respiratory infection: Is it an innocent bystander?

PURPOSE: Acute respiratory infection (ARI) is one of the major attributing factors of under-five mortality and morbidity all over the world. Viruses are the most common cause of ARI. Due to the availability of molecular techniques, new viruses are getting isolated from children with ARI. With the above background, the present study was conducted to enlighten on the pathogenic role of human bocavirus (HBoV) in children with ARI. METHODOLOGY: This retrospective study was conducted over a period of >3 years duration. The clinical and laboratory data of the patients with signs and symptoms of ARI were retrieved and analyzed. Clinical profiles and outcome of the patients detected of having HBoV mono or co-infections were further analyzed in details. RESULTS: A total of 237 respiratory samples were subjected to respiratory panel by fast track diagnosis (FTD) multiplex polymerase chain reaction (multiplex PCR), of which 10 samples (mono-infection â€‹= â€‹4) were detected with the presence of HBoV. The clinical details of 8 cases were studied in details (details of rest 2 cases were missing). All the children were less than 3 years of age, with different co-morbid conditions such as low birth weight (n â€‹= â€‹4), cholestatic jaundice (n â€‹= â€‹1), operated case of congenital diaphragmatic hernia (n â€‹= â€‹1), pancytopenia (n â€‹= â€‹1), and primary immune deficiency (n â€‹= â€‹1). Their clinical course did not improve following antibiotic administration, 2 succumbed to death while the rest 6 cases were discharged. CONCLUSION: The present study highlights the fact that HBoV may not be an innocent bystander in the childhood ARI. Larger studies employing appropriate diagnostic modalities are needed to emboss it as a true pathogen and not merely a bystander.

Human bocavirus respiratory infection: Tracing the path from viral replication and virus-cell interactions to diagnostic methods.

Human bocaviruses were first described between 2005 and 2010, identified in respiratory and enteric tract samples of children. Screening studies have shown worldwide distribution. Based on phylogenetic analysis, they were classified into four genotypes (HBoV1-4). From a clinical perspective, human bocavirus 1 (HBoV1) is considered the most relevant, since it can cause upper and lower acute respiratory tract infection, mainly in infants, including common cold, bronchiolitis, and pneumonia, as well as wheezing in susceptible patients. However, the specific processes leading to structural, biochemical, and functional changes resulting in the different clinical presentations have not been elucidated yet. This review surveys the interactions between the virus and target cells that can potentially explain disease-causing mechanisms. It also summarises the clinical phenotype of cases, stressing the role of HBoV1 as an aetiological agent of lower acute respiratory infection in infants, together with laboratory tests for detection and diagnosis. By exploring the current knowledge on the epidemiology of HBoV1, insights into the complex scenario of paediatric respiratory infections are presented, as well as the potential effects that changes in the circulation can have on the dynamics of respiratory agents, spotlighting the benefits of comprehensively increase insights into incidence, interrelationships with co-circulating agents and potential control of HBoV1.

HBoV-1: virus structure, genomic features, life cycle, pathogenesis, epidemiology, diagnosis and clinical manifestations.

The single-stranded DNA virus known as human bocavirus 1 (HBoV-1) is an icosahedral, linear member of the Parvoviridae family. In 2005, it was discovered in nasopharyngeal samples taken from kids who had respiratory tract illnesses. The HBoV genome is 4.7-5.7 kb in total length. The HBoV genome comprises three open-reading frames (ORF1, ORF2, and ORF3) that express structural proteins (VP1, VP2, and VP3), viral non-coding RNA, and non-structural proteins (NS1, NS1-70, NS2, NS3, and NP1) (BocaSR). The NS1 and NP1 are crucial for viral DNA replication and are substantially conserved proteins. Replication of the HBoV-1 genome in non-dividing, polarized airway epithelial cells. In vitro, HBoV-1 infects human airway epithelial cells that are strongly differentiated or polarized. Young children who have HBoV-1 are at risk for developing a wide range of respiratory illnesses, such as the common cold, acute otitis media, pneumonia, and bronchiolitis. The most common clinical symptoms are wheezing, coughing, dyspnea, and rhinorrhea. After infection, HBoV-1 DNA can continue to be present in airway secretions for months. The prevalence of coinfections is considerable, and the clinical symptoms can be more severe than those linked to mono-infections. HBoV-1 is frequently detected in combination with other pathogens in various reports. The fecal-oral and respiratory pathways are more likely to be used for HBoV-1 transmission. HBoV-1 is endemic; it tends to peak in the winter and spring. This Review summarizes the knowledge on HBoV-1.

Molecular Profiling of Inflammatory Mediators in Human Respiratory Syncytial Virus and Human Bocavirus Infection.

Infections due to human respiratory syncytial virus (HRSV) and human bocavirus (HBoV) can mediate the release of several pro-inflammatory cytokines such as IL-6, IL-8, and TNF-α, which are usually associated with disease severity in children. In this study, the change in the expression profile of cytokines and chemokines were determined during HRSV, HBoV, and HRSV coinfection with HBoV in 75 nasopharyngeal aspirates (NPAs) samples, positive real-time reverse transcriptase PCR Assay (rRT-PCR) for HRSV (n = 36), HBoV (n = 23) infection alone or HRSV coinfection with HBoV (n = 16). The samples were collected from hospitalized children. qPCR-based detection revealed that the levels of IL-6, IL-8, IL-10, IL-13, IL-33, and G-CSF were significantly (p < 0.05) greater in patients than in controls. IL-4, IL-17, GM-CSF, and CCL-5 were significantly elevated in children with HRSV coinfection with HBoV than in other groups (p < 0.05). TNF-α, IL-6, IL-8, IL-10, IL-13, and IL-33 in children with HRSV were significantly increased in severe infections compared to mild infections. Whereas, IL-10, IL-13, and IL-33 were significantly increased in severe infection in compared a mild infection in children with HBoV. Further large-scale investigations involving isolates are needed to enhance our knowledge of the association between viral infections and cytokine expression patterns during the different stages of HRSV and HBoV infection.

Epidemiologic and Clinical Characteristics of Human Bocavirus Infection in Children with or without Acute Gastroenteritis in Acre, Northern Brazil.

Human bocavirus (HBoV) is an emerging virus detected around the world that may be associated with cases of acute gastroenteritis (AGE). However, its contribution to AGE has not been elucidated. This study aimed to describe the frequency, clinical features, and HBoV species circulation in children up to 5 years with or without AGE symptoms in Acre, Northern Brazil. A total of 480 stool samples were collected between January and December 2012. Fecal samples were used for extraction, nested PCR amplification, and sequencing for genotyping. Statistical analysis was applied to verify the association between epidemiological and clinical characteristics. Overall, HBoV-positivity was 10% (48/480), with HBoV-positive rates of 8.4% (19/226) and 11.4% (29/254) recorded in diarrheic and non-diarrheic children, respectively. The most affected children were in the age group ranging between 7 and 24 months (50%). HBoV infection was more frequent in children who live in urban areas (85.4%), use water from public networks (56.2%), and live with adequate sewage facilities (50%). Co-detection with other enteric viruses was 16.7% (8/48) and the most prevalent coinfection was RVA+ HBoV (50%, 4/8). HBoV-1 was the most frequent species detected in diarrheic and non-diarrheic children, responsible for 43.8% (21/48) of cases, followed by HBoV-3 (29.2%, 14/48) and HBoV-2 (25%, 12/48). In this study, HBoV infection was not always associated with AGE, as most HBoV cases belonged to the non-diarrheal group. Future studies are warranted in order to determine the role of HBoV in causing acute diarrhea disease.

The Burden of Human Bocavirus 1 in Hospitalized Children With Respiratory Tract Infections.

BACKGROUND: Human bocavirus 1 (HBoV1) is frequently codetected with other viruses, and detected in asymptomatic children. Thus, the burden of HBoV1 respiratory tract infections (RTI) has been unknown. Using HBoV1-mRNA to indicate true HBoV1 RTI, we assessed the burden of HBoV1 in hospitalized children and the impact of viral codetections, compared with respiratory syncytial virus (RSV). METHODS: Over 11 years, we enrolled 4879 children <16 years old admitted with RTI. Nasopharyngeal aspirates were analyzed with polymerase chain reaction for HBoV1-DNA, HBoV1-mRNA, and 19 other pathogens. RESULTS: HBoV1-mRNA was detected in 2.7% (130/4850) samples, modestly peaking in autumn and winter. Forty-three percent with HBoV1 mRNA were 12-17 months old, and only 5% were <6 months old. A total of 73.8% had viral codetections. It was more likely to detect HBoV1-mRNA if HBoV1-DNA was detected alone (odds ratio [OR]: 3.9, 95% confidence interval [CI]: 1.7-8.9) or with 1 viral codetection (OR: 1.9, 95% CI: 1.1-3.3), compared to ≥2 codetections. Codetection of severe viruses like RSV had lower odds for HBoV1-mRNA (OR: 0.34, 95% CI: 0.19-0.61). The yearly lower RTI hospitalization rate per 1000 children <5 years was 0.7 for HBoV1-mRNA and 8.7 for RSV. CONCLUSIONS: True HBoV1 RTI is most likely when HBoV1-DNA is detected alone, or with 1 codetected virus. Hospitalization due to HBoV1 LRTI is 10-12 times less common than RSV.

Human bocavirus 1 epidemiology in children in relation to virus load and codetection.

AIM: Human bocavirus 1 (HBoV1) has been associated with respiratory tract infections in children. We aimed at retrospectively describing patient characteristics, seasonality, pre-existing medical conditions, codetections, clinical manifestations and complications of HBoV1 infection in relation to viral load in the child population in Stockholm, with the overarching aim of elucidating the clinical significance of HBoV1. METHODS: We included all hospitalised children 0-17 years testing positive for HBoV1 by real-time polymerase chain reaction on nasopharyngeal aspirates 1 July 2008-30 June 2019. Patients with HBoV1 single detection, high viral load expressed as an HBoV1-DNA cycle threshold (Ct) < 25, or both, were separately analysed. We retrieved information on pre-existing conditions and clinical course from the medical records. RESULTS: We found 768 episodes in 727 children, 496 (64.6%) male and 441 (60.7%) previously healthy. The median age was 17.6 months. Most (476/768, 62.0%) episodes occurred during December-March. HBoV1 was in 549 episodes (71.5%) codetected with other viruses. Ct < 25 was independently associated with young age, single detection of HBoV1 and presentation early in the epidemic season. We saw few differences in clinical manifestations between the subgroups. CONCLUSION: Our findings are consistent with primary HBoV1 infection causing mild-to-severe respiratory tract manifestations in young children.

Human bocavirus 1 and 2 genotype-specific antibodies for rapid antigen testing in pediatric patients with acute respiratory infections.

BACKGROUND: Previous serological studies of human bocavirus (HBoV) 1 could not exclude cross-reactivity with the other three HBoVs, particularly HBoV2. METHODS: To search for genotype-specific antibodies against HBoV1 and HBoV2, the divergent regions (DRs) located on the major capsid protein VP3 were defined through viral amino acid alignment and structure prediction. DR-deduced peptides were used as antigens to harvest corresponding anti-DR rabbit sera. To determine their genotype specificities for HBoV1 and HBoV2, these sera samples were used as antibodies against the antigens VP3 of HBoV1 and HBoV2 (expressed in Escherichia coli) in western blotting (WB), enzyme-linked immunosorbent assay (ELISA), and bio-layer interferometry (BLI) assays. Subsequently, the antibodies were evaluated with clinical specimens from pediatric patients with acute respiratory tract infection by indirect immunofluorescence assay (IFA). RESULTS: There were four DRs (DR1-4) located on VP3 with different secondary and tertiary structures between HBoV1 and HBoV2. Regarding the reactivity with VP3 of HBoV1 or HBoV2 in WB and ELISA, high intra-genotype cross-reactivity of anti-HBoV1 or HBoV2 DR1, DR3, and DR4, but not anti-DR2, was observed. Genotype-specific binding capacity of anti-DR2 sera was confirmed by BLI and IFA, in which only anti-HBoV1 DR2 antibody reacted with HBoV1-positive respiratory specimens. CONCLUSION: Antibodies against DR2, located on VP3 of HBoV1 or HBoV2, were genotype specific for HBoV1 and HBoV2, respectively.

Human Bocavirus in children with acute gastroenteritis in Piedmont, Italy.

BACKGROUND: Gastroenteritis is a common disease in children, characterized by diarrhea, vomiting, abdominal pain, and fever. Co-detection of human Bocavirus (HBoV) with other gastroenteric viruses was reported a lot in patients with acute gastroenteritis. METHODS: This paper presents the real-time RT-PCR Taqman assay for the detection and quantification of HBoV for clinical fecal samples collected from hospitalized children with acute gastroenteritis in Piedmont. RESULTS: All fecal specimens were tested for the presence of HBoV with specific primers and probe. A total of 17 out of 123 (13.92%) episodes of acute gastroenteritis were associated with HBoV genomic detection with median viral load 6864.75±19784.79 genomes/mg fecal specimens. Among the 17 HBoV-positive cases, 11 were also positive for other viral pathogens, including Rotavirus (N.=2), astrovirus (N.=1), norovirus GII (N.=6), norovirus GI (N.=2). Two cases were positive for more than one virus including norovirus GII and norovirus GI (N.=1) and Rotavirus, sapovirus and astrovirus (N.=1). A higher detection of HBoV infections was observed in winter, and peaking in February. CONCLUSIONS: Although HBoV is suspected to be responsible for gastroenteritis in children, our data showed that this association was uncertain since no difference was observed in term of viral load in the group with single infection of HBoV and group of coinfections with other viral agent.

Molecular characterization and clinical impact of human bocavirus at a tertiary hospital in Barcelona (Catalonia, Spain) during the 2014-2017 seasons.

PURPOSE: The aim was to describe the prevalence, molecular epidemiology and clinical manifestations of human bocavirus (HBoV) in patients attended at a tertiary hospital in Barcelona, Spain. METHODS: From October 2014 to May 2017, respiratory specimens from paediatric patients were collected for respiratory viruses' laboratory-confirmation. Phylogenetic analyses from partial VP1 sequences were performed from all HBoV laboratory-confirmed specimens. Clinical features were retrospectively studied. RESULTS: 178/10271 cases were HBoV laboratory-confirmed. The median age was 1.53 (IQR 1.0-2.3). Co-detection was highly reported (136; 76%). All viruses belonged into HBoV1 genotype but one into HBoV2. Non-reported mutations were observed and two sites were suggestive to be under negative selection. 61% (109/178) cases had lower RTI (LRTI), of whom 84 had co-detections (77%) and 76 had comorbidities (70%). LRTI was the cause of hospitalization in 85 out of 109 cases (78%), and no differences were found regarding severity factors during hospitalization between co- and single-detections, except for median length of respiratory support, which was longer in cases with co-detections. CONCLUSIONS: Close monitoring of predominant HBoV1 showed a high similarity between viruses. The presence of comorbidities might explain the high prevalence of LRTI. Symptomatology in HBoV single-detected cases suggest that HBoV is a true pathogen.

Plastic Bronchitis of Human Bocavirus 1 Detected by Comprehensive Polymerase Chain Reaction of Mucus Casts.

Plastic bronchitis (PB) is a rare and severe respiratory disease characterized by the formation of branching mucus casts, resulting in airway obstruction. PB can be divided into two types. Type 1 PB is primarily caused by inflammatory casts that result from allergic diseases. In type 2 PB, mucinous casts are produced in association with congenital heart disease. PB is also associated with viral respiratory infections, particularly the influenza A (H1N1) pdm09 virus, which is the most common pathogen affecting pediatric patients. Herein, we report a case of severe type 1 PB caused by human bocavirus (HBoV)1 in a child. Multiplex polymerase chain reaction (PCR) of a nasopharyngeal swab revealed the presence of respiratory syncytial virus and human parainfluenza virus 3. However, no viruses other than HBoV1 were detected in mucus casts by real-time PCR. Consequently, we suggest that HBoV can cause PB in pediatric patients, and direct and comprehensive PCR of bronchial casts may be useful for identifying the etiologic agents.