Genetic and epidemiological characteristics of the spread of human metapneumovirus in Bulgaria, 2023–2025

Introduction Human metapneumovirus (hMPV) can cause severe respiratory illnesses such as bronchitis, bronchiolitis, and pneumonia, particularly in young children, older adults, and individuals with weakened immune systems. This study aimed to assess hMPV circulation patterns, including seasonal and demographic distribution, track genetic evolution and mutations in surface proteins, and explore associations with clinical severity. Methods From early 2023 to November 2025, Bulgaria’s National Reference Laboratory tested nasopharyngeal samples from 6,958 patients with respiratory symptoms using an in-house multiplex real-time PCR method that detected 11 viruses. Positive hMPV samples were sequenced and analyzed phylogenetically. Results A study conducted 3 years after the COVID-19 pandemic found that 1.4% (97 patients) tested positive for hMPV, with most being mono-infected (85.6%) and others co-infected, primarily with rhinovirus and adenovirus. The infection rate was highest in children under 5 (2.8%), especially infants and toddlers (2.2%), compared to 0.4% in those over 65 ( p < 0.05). Common symptoms in 56% of hMPV-positive patients included cough (50%), fever (38%), and rhinitis (21.2%). Among 94 patients with respiratory involvement, 34% had upper respiratory tract infections (URTs), and 55% had lower respiratory tract infections (LRTs), with bronchiolitis (46%) and pneumonia (30.7%) being common. Hospitalized patients had a 19% rate of severe acute respiratory infection (SARI), higher than the 7.3% for other viruses. Sequencing revealed genotypes A and B in similar proportions, with subclade B2 being the most common at 41.3%. However, in 2025, subclade A2b2 represented 52% of the samples. The A2b2 subclade in children under 5 showed a higher incidence of lower respiratory complications ( p < 0.05), while genotype B was linked to an increased risk of SARI at higher viral loads ( p < 0.05). Conclusion The findings confirm that hMPV is a significant respiratory pathogen for young children, showing substantial genetic diversity in Bulgaria, with lineage A2b2 becoming more prevalent in 2025. Host age and viral genetics may influence disease presentation. These results highlight the clinical burden of hMPV and emphasize the need for preventive strategies, including vaccine development for high-risk children

Bronchiolitis · Human metapneumovirus · Pneumonia · Respiratory system · Respiratory tract infections · Rhinovirus · Subclade · Delphi Technique in Research · Pediatric health and respiratory diseases · Respiratory viral infections research · Epidemiology