Katherine E. Arden

Frequent detection of human rhinoviruses, paramyxoviruses, coronaviruses, and bocavirus during acute respiratory tract infections.

Viruses are the major cause of pediatric acute respiratory tract infection (ARTI) and yet many suspected cases of infection remain uncharacterized. We employed 17 PCR assays and retrospectively screened 315 specimens selected by season from a predominantly pediatric hospital‐based population. Before the Brisbane respiratory virus research study commenced, one or more predominantly viral pathogens had been detected in 15.2% (n = 48) of all specimens. The Brisbane study made an additional 206 viral detections, resulting in the identification of a microbe in 67.0% of specimens. After our study, the majority of microbes detected were RNA viruses (89.9%). Overall, human rhinoviruses (HRVs) were the most frequently identified target (n = 140) followed by human adenoviruses (HAdVs; n = 25), human metapneumovirus (HMPV; n = 18), human bocavirus (HBoV; n = 15), human respiratory syncytial virus (HRSV; n = 12), human coronaviruses (HCoVs; n = 11), and human herpesvirus‐6 (n = 11). HRVs were the sole microbe detected in 37.8% (n = 31) of patients with suspected lower respiratory tract infection (LRTI). Genotyping of the HRV VP4/VP2 region resulted in a proposed subdivision of HRV type A into sublineages A1 and A2. Most of the genotyped HAdV strains were found to be type C. This study describes the high microbial burden imposed by HRVs, HMPV, HRSV, HCoVs, and the newly identified virus, HBoV on a predominantly paediatric hospital population with suspected acute respiratory tract infections and proposes a new formulation of viral targets for future diagnostic research studies. J. Med. Virol. 78:1232–1240, 2006.

Evidence of human coronavirus HKU1 and human bocavirus in Australian children

Abstract Undiagnosed cases of respiratory tract disease suspected of an infectious aetiology peak during the winter months. Since studies applying molecular diagnostic assays usually report reductions in the number of undiagnosed cases of infectious disease compared to traditional techniques, we applied PCR assays to investigate the role of two recently described viruses, namely human coronavirus (HCoV) HKU1 and human bocavirus (HBoV), in a hospital-based paediatric population. Both viruses were found among Australia children with upper or lower respiratory tract disease during the autumn and winter of 2004, contributing to 21.1% of all microbial diagnoses, with individual incidences of 3.1% (HCoV-HKU1) and 5.6% (HBoV) among 324 specimens. HBoV was found to coincide with another virus in more than half of all instances and displayed a single genetic lineage, whilst HCoV-HKU1 was more likely to occur in the absence of another microbe and strains could be divided into two genetic lineages which we propose be termed HCoV-HKU1 type A and type B. Children under the age of 2 years were most at risk of infection by these viruses which contribute significantly to the microbial burden among patients with respiratory tract disease during the colder months.

New human coronavirus, HCoV-NL63, associated with severe lower respiratory tract disease in Australia

A new human coronavirus, HCoV‐NL63, was associated recently with bronchiolitis. The current study aimed to examine retrospectively stored specimens for the presence of HCoV‐NL63 using nested RT‐PCR assays targeting the 1a and 1b genes. The study population was composed of patients with acute respiratory disease warranting presentation to Queensland hospitals. HCoV‐NL63 was detected in the nasopharyngeal aspirates (NPA) of 16 of 840 specimens representing 766 patients (2%). HCoV‐NL63 positive individuals were diagnosed most commonly with lower respiratory tract (LRT) disease (81%). The clinical diagnosis was commonly supported by an abnormal chest X‐ray (56%) together with respiratory distress (50%), wheeze (44%), and râles (25%) on first presentation with HCoV‐NL63 infection. All patients positive for HCoV‐NL63 required admission to hospital. Among 38% of HCoV‐NL63 positive specimens a second pathogen was detected. Sequencing of amplicon from gene 1b revealed more than 99% nucleotide homology with the viral type strains while sequencing amplicon from gene 1a permitted the grouping of viral strains. It was shown that HCoV‐NL63 is associated with severe LRT disease in an Australian hospital setting during the cooler months of the year. We propose that HCoV‐NL63 is a global and seasonal pathogen of both children and adults associated with severe LRT illness. J. Med. Virol. 75:455–462, 2005.

Do rhinoviruses reduce the probability of viral co-detection during acute respiratory tract infections?

Abstract Background Human rhinoviruses (HRVs) are often concurrently detected with other viruses found in the respiratory tract because of the high total number of HRV infections occurring throughout the year. This feature has previously relegated HRVs to being considered passengers in acute respiratory infections. HRVs remain poorly characterized and are seldom included as a target in diagnostic panels despite their pathogenic potential, infection-associated healthcare expenditure and relatively unmoderated elicitation of an antiviral state. Objectives To test the hypothesis that respiratory viruses are proportionately more or less likely to co-occur, particularly the HRVs. Study design Retrospective PCR-based analyses of 1247 specimens for 17 viruses, including HRV strains, identified 131 specimens containing two or more targets. We investigated the proportions of co-detections and compared the proportion of upper vs. lower respiratory tract presentations in the HRV positive group. Both univariate contingency table and multivariate logistic regression analyses were conducted to identify trends of association among the viruses present in co-detections. Results Many of the co-detections occurred in patterns. In particular, HRV detection was associated with a reduced probability of detecting human adenoviruses, coronaviruses, bocavirus, metapneumovirus, respiratory syncytial virus, parainfluenza virus, influenza A virus, and the polyomaviruses KIPyV and WUPyV (p ≤0.05). No single HRV species nor cluster of particular strains predominated. Conclusions HRVs were proportionately under-represented among viral co-detections. For some period, HRVs may render the host less likely to be infected by other viruses.

A novel mutation in ferroportin1 is associated with haemochromatosis in a Solomon Islands patient

Background: A severe form of iron overload with the clinicopathological features of haemochromatosis inherited in an autosomal dominant manner has been described in the Solomon Islands. The genetic basis of the disorder has not been identified. The disorder has similarities to type 4 haemochromatosis, which is caused by mutations in ferroportin1. Aims: The aims of this study were to identify the genetic basis of iron overload in a patient from the Solomon Islands. Patient and methods: Genomic DNA was isolated from peripheral blood leucocytes of a Solomon Islands man with severe iron overload. The entire coding region and splice sites of the ferroportin1 gene was sequenced. Results and conclusions: A novel missense mutation (431A>C; N144T) was identified in exon 5 of the ferroportin1 gene. A novel restriction endonuclease based assay which identifies both the N144T and N144H mutations was developed which will simplify the diagnosis and screening of patients for iron overload in the Solomon Islands and other populations. This is the first identified mutation associated with haemochromatosis in the Solomon Islands population.

Newly identified human rhinoviruses: molecular methods heat up the cold viruses.

Human rhinovirus (HRV) infections cause at least 70% of virus‐related wheezing exacerbations and cold and flu‐like illnesses. They are associated with otitis media, sinusitis and pneumonia. Annually, the economic impact of HRV infections costs billions in healthcare and lost productivity. Since 1987, 100 officially recognised HRV serotypes reside in two genetically distinct species; HRV A and HRV B, within the genus Enterovirus, family Picornaviridae. Sequencing of their ∼7kb genomes was finalised in 2009. Since 1999, many globally circulating, molecularly‐defined ‘strains’, perhaps equivalent to novel serotypes, have been discovered but remain uncharacterised. Many of these currently unculturable strains have been assigned to a proposed new species, HRV C although confusion exists over the membership of the species. There has not been sufficient sampling to ensure the identification of all strains and no consensus criteria exist to define whether clinical HRV detections are best described as a distinct strain or a closely related variant of a previously identified strain (or serotype). We cannot yet robustly identify patterns in the circulation of newly identified HRVs (niHRVs) or the full range of associated illnesses and more data are required. Many questions arise from this new found diversity: what drives the development of so many distinct viruses compared to other species of RNA viruses? What role does recombination play in generating this diversity? Are there species‐ or strain‐specific circulation patterns and clinical outcomes? Are divergent strains sensitive to existing capsid‐binding antivirals? This update reviews the findings that trigger these and other questions arising during the current cycle of intense rhinovirus discovery. Copyright

Genetic Diversity of Human Metapneumovirus over 4 Consecutive Years in Australia

The molecular epidemiologic profile of human metapneumovirus (hMPV) infection has likely been skewed toward certain genetic subtypes because of assay-design issues, and no comprehensive studies have been conducted to date. Here, reverse-transcription polymerase chain reaction was used to screen 10,319 specimens from patients presenting to hospitals with suspected respiratory tract infections during 2001-2004. After analysis of 727 Australian hMPV strains, 640 were assigned to 1 of 4 previously described subtypes. hMPV was the most common pathogen detected, and subtype B1 was the most common lineage. Concurrent, annual circulation of all 4 hMPV subtypes in our study population was common, with a single, usually different hMPV subtype predominating in each year.

Newly Identified Respiratory Viruses in Children With Asthma Exacerbation not Requiring Admission to Hospital

There are few data describing the comprehensive identification in and influence of newly identified respiratory viruses on asthma exacerbations. Most studies focus on inpatients. In this preliminary study, the point prevalence and the associations of picornavirus species described recently and human bocavirus (HBoV) with the recovery from exacerbations in non‐hospitalized asthmatic children (median age 5.1 years) were examined. Human rhinoviruses (HRVs) were present in 52.6% of specimens, HBoV‐1 was in 7.7%. Viral co‐detections occurred in 25.6% of children and were associated (P = 0.04) with lower asthma quality of life scores upon presentation than were single viral detections. The undifferentiated presence or absence of virus did not influence the severity of asthma or recovery however when virus species were examined individually, specific clinical associations emerged. HRV species C (HRV‐Cs) were the viruses most frequently detected as single virus detections. Among 41 genotyped HRVs, more HRV‐Cs (n = 23) were identified than HRV‐As (n = 16) however HRV‐A detection was associated (P = 0.01) with worse asthma symptoms and cough for longer than was HRV‐C detection. Larger, PCR‐based studies are required to elucidate further the true impact of HRV species in childhood asthma exacerbations of both hospitalized and non‐hospitalized cohorts. J. Med. Virol. 82:1458–1461, 2010.

Molecular characterization and distinguishing features of a novel human rhinovirus (HRV) C, HRVC-QCE, detected in children with fever, cough and wheeze during 2003

Abstract Background Human rhinoviruses (HRVs) are associated with more acute respiratory tract infections than any other viral group yet we know little about viral diversity, epidemiology or clinical outcome resulting from infection by strains, in particular the recently identified HRVs. Objectives To determine whether HRVC-QCE was a distinct HRV-C strain, by determining its genome and prevalence, by cataloguing genomic features for strain discrimination and by observing clinical features in positive patients. Study design Novel real-time RT-PCRs and retrospective chart reviews were used to investigate a well-defined population of 1247 specimen extracts to observe the prevalence and the clinical features of each HRV-QCE positive case from an in- and out-patient pediatric, hospital-based population during 2003. An objective illness severity score was determined for each HRVC-QCE positive patient. Results Differences in overall polyprotein and VP1 binding pocket residues and the predicted presence of a cis-acting replication element in 1B defined HRVC-QCE as a novel HRV-C strain. Twelve additional HRVC-QCE detections (1.0% prevalence) occurred among infants and toddlers (1–24 months) suffering mild to moderate illness, including fever and cough, who were often hospitalized. HRVC-QCE was frequently detected in the absence of another virus and was the only virus detected in three (23% of HRVC-QCE positives) children with asthma exacerbation and in two (15%) toddlers with febrile convulsion. Conclusions HRVC-QCE is a newly identified, genetically distinct HRV strain detected in hospitalized children with a range of clinical features. HRV strains should be independently considered to ensure we do not overestimate the HRVs in asymptomatic illness.

Community-Wide, Contemporaneous Circulation of a Broad Spectrum of Human Rhinoviruses in Healthy Australian Preschool-Aged Children During a 12-Month Period

Abstract Human rhinovirus (HRV) replication triggers exacerbation of asthma and causes most acute respiratory illnesses (ARIs), which may manifest as influenza-like illness. The recent assignment of 60 previously unknown HRV types to a third HRV species, Human rhinovirus C, raised questions about the prevalence of these picornavirus types in the community, the extent of HRV diversity at a single site, and whether the HRVs have an equally diverse clinical impact on their hosts. We quantified HRV diversity, and there was no clinical impact attributable to HRV species and genotypes among a community population of preschool-aged children with ARI who provided respiratory samples during 2003. All HRV species were represented among 138 children with ARI, and 74 distinct HRV types were cocirculating. Fever accompanied 32.8% of HRV-positive ARI cases. HRVs were less likely than DNA viruses to be codetected with another virus, suggesting virus interference at the community level, demonstrated by the inverse correlation between influenza virus detection and HRV detection.