Multi-year Study of Human Metapneumovirus Infection

1. Multi-year Study of Human Metapneumovirus Infection at a Large US Midwestern Medical Referral Center Abstract Acknowledgements Discussion Introduction Results Materials & Methods Conclusion Gregory C. Gray, MD, MPH,1 Ana W. Capuano, MPS,1 Sharon F. Setterquist,1 Dean D. Erdman, DrPH,2 Newell D. Nobbs,3 Yacine Abed, PhD,4 Gary V. Doern, PhD,5 Sarah E. Starks, MS,1 and Guy Boivin, MD, MSC, FECPC4 1Center for Emerging Infectious Diseases, Department of Epidemiology, University of Iowa College of Public Health, Iowa City, IA 2Respiratory and Enteric Viruses Branch, Center for Disease Control and Prevention, Atlanta, GA 3Hospital Information Systems, University of Iowa College of Public Health, Iowa City, IA 4Centre Hospitalier Universitaire de Québec, Québec City, Québec, Canada 5Section Director, Clinical Microbiology, University of Iowa Hospital and Clinics, Iowa City, IA Using RT-PCR, we retrospectively screened 1294 archived clinical specimens for human metapneumovirus. The specimens were submitted to a large US Midwest referral hospital. Testing by RT-PCR identified 34 (2.6%) hMPV positive samples. Among these, 21 (62%) were culture positive and available for genetic typing. A previously considered rare genotype of hMPV, B1, was the most common single genotype identified, comprising 9 (43%) of the 21 isolates. An analysis of available demographic and clinical data from these patients revealed that hMPV infection was associated with patient age as well as severe morbidity and an immunocompromised state. Multivariate logistic regression modeling identified patients aged 0.4 to 9 yrs (OR=8.9; 95% CI=2.0-38.5) and those under intensive care (OR=3.2; 95%CI=1.1-8.7) as more likely to have hMPV infection than their peers. • The 34 hMPV+ specimens were obtained from patients with a wide array of clinical conditions, many immunocompromised. 4 had bacterial infections, 4 had other viral infections, and one had a fungal infection. These coinfections support the important potential role of hMPV as a coinfecting pathogen. • The increased risk and the increasing trend in hMPV prevalence (Fig. 1) up to age 6 suggests hMPV surveillance and preventive measures may be most efficiently focused upon patients less than 6 years of age. • hMPV infections may be more common among intensive care unit patients. • A relative high proportion of isolates (43% of 21 sequenced) were classified in the B1 genome subgroup (Fig. 2). • Diagnostics and genetic characterization- • 34 (2.6%) of the 1294 specimens available for study were confirmed to be RT-PCR+. • 21 (62%) of the RT-PCR+ specimens grew hMPV virus in LLC-MK2 cell cultures. • Clinical features- • Among the 34 hMPV+ patients: • 7 presented with respiratory failure. • 27 were hospitalized when cultured, with the median hospital stay of 8 days. • Frequent diagnoses included pneumonia, bronchiolitis, and nonspecific acute upper respiratory infection. • Among the five hMPV genotype B isolates, 4 (80%) were of subtype B1. • Risk factor modeling- • The median patient age was 5.3 years, 52% were male and 66.8% were Caucasian. 83.5% of the specimens were collected during the months of December through April. • Most (61.8%) of the hMPV+ specimens were from patients with ages of 0.4 to 9 years. A significant increasing trend of hMPV+ prevalence by age was noticed in infants up to 6 years old (Fig. 1). • Multivariable modeling- • Only age group and patient group were associated with hMPV+ specimens (Table 1). • Children between 0.4 and 9 years of age had 8.9 times the odds of having hMPV infection compared with infants 0.4 years old or younger (Table 1). • Intensive care unit patients had a higher odds of being hMPV+ as compared to outpatients. • Phylogenetic analyses of hMPV strains- • There was a high prevalence of subtypes B1 (9 isolates) and A2 (6 isolates) (Fig. 2). Since the human metapneumovirus (hMPV) was first reported in 2001, much progress has been made in its detection, in understanding its genetic diversity and in documenting its varied clinical presentation. However, because of its recent identification, and infrequency of infection, few multi-year, epidemiologic studies of hMPV infection have been reported. By linking demographic and clinical data to a large, 3-year, viral culture specimen repository, we sought to retrospectively describe hMPV infections among patients evaluated by a large US Midwestern referral laboratory. Our retrospective study of respiratory specimens collected from 1294 patients by a large tertiary Midwest US hospital revealed that 34 (2.6%) had evidence of hMPV infection. Many of the hMPV+ patients had severe morbidity and were immunocompromised. Infection occurred among all age groups, but those aged 0.4 to 9 yrs were at greatest risk. Among patient groups, those in an intensive care had 3 times the odds of being infected with hMPV. Finally, a previously considered rare subtype of hMPV (B1) was found to be the most prevalent, comprising 9 (43%) of the genotyped isolates. Fig. 1. Distribution of hMPV RT-PCR+ specimen from patients with ages 0 to 9 yrs old. • Specimen collection and Clinical Data- • From October 1, 2001 to May 18, 2004, 1500 archived respiratory specimens were collected from the Clinical Microbiology Laboratory at the University of Iowa Hospitals • and Clinics. • Specimens originated from inpatient wards, intensive care units, and hospital affiliated outpatient clinics. • Demographic data and clinical characteristics were retrieved using a relational database. • RT-PCR- • Extracted RNA was screened with a one-step RT-PCR F-gene procedure. • Screened specimens that yielded bands within 50bp of the expected 347bp product • were further tested with additional primer sets. • hMPV isolation- • RT-PCR+ specimens were further studied with LLC-MK2 shell-vial culture. • Sequencing- • Amplification products of a 800-1000 bp region of the hMPV G protein gene were electrophoresed, purified, and then sequenced in both directions. • Phylogenetic analysis- • Alignments of the partial nucleotide sequences of the hMPV G-gene were generated. • Prototype sequences of different hMPV genotypes and subtypes were included. • Phylogenetic trees were computed by the neighbor joining method. • Statistical analysis- • Binomial confidence intervals, and unadjusted and adjusted odds ratios using logistic regression were computed. • The final multivariable model used a saturated model and manual backwards elimination. Table 1. Prevalence and odds ratio of RT-PCR positivity for human metapneumovirus by risk factor, without duplicates. The findings and conclusions on this report are those of the authors and do not necessarily represent the views of the funding agency. This work was funded by the National Institute of Allergy and Infectious Diseases under proposal R03AI054570-01A1. We thank Mark LeBeck and Troy McCarthy of the Center for Emerging Infectious Diseases for their help with hMPV sequencing and the phylogenetic analysis. Fig. 2. Neighbor joining tree computed from alignment of partial nucleotide sequences of the hMPV G gene.