Blood and Respiratory TTV Load, Anellovirus Species Distribution, and Compartment Overlap in Acute Respiratory Viral Infections

ABSTRACT

Human anelloviruses are ubiquitous members of the commensal virome, and blood Torquetenovirus (TTV) load has emerged as a candidate marker of host immune status. Their behavior during acute respiratory viral infections across blood and respiratory compartments remains poorly defined. To assess TTV DNA load by quantitative PCR, anellovirus species distribution by next‐generation sequencing, soluble immune markers, and blood‐respiratory genomic overlap of anelloviruses in hospitalized patients with acute respiratory viral infection. Study design: A single‐center observational study was performed on leftover specimens from patients admitted to the University Hospital of Varese, Italy, between September 2024 and March 2025. We collected 37 respiratory specimens and 9 plasma samples. Respiratory viruses were identified by multiplex real‐time PCR. TTV DNA was quantified in blood and respiratory samples by an in‐house universal TaqMan assay targeting the untranslated region; this assay provides a total TTV DNA measurement. Anellovirus species composition was characterized separately by amplicon‐based next‐generation sequencing. Plasma cytokines and adhesion molecules were measured by automated microfluidic immunoassay. In five paired blood‐respiratory cases, reconstructed anellovirus genomes were compared at the whole‐genome level. Total TTV load differed by infecting virus, with significantly higher levels in human rhinovirus (HRV) than influenza (FLU) or respiratory syncytial virus (RSV) in both blood and respiratory samples. IFN‐β was higher in FLU than RSV, whereas ICAM‐1 was higher in HRV than RSV. Age had a stronger effect than sex: children aged 0‐10 years had higher blood and respiratory TTV loads and higher IFN‐α, IFN‐β, and ICAM‐1, whereas adults aged 34–90 years showed greater representation of TTV9 and TTV24. No significant association was found between the number of detected anellovirus species and TTV DNA load. Whole‐genome comparison showed partial overlap of blood‐ and respiratory‐derived anelloviruses without compartment‐specific clustering. In acute respiratory viral infections, TTV DNA load and age‐related differences in anellovirus community composition provided information distinct from simple anellovirus species counts. HRV infection was associated with the highest total TTV burden, and blood and respiratory compartments shared part of the same anellovirus population.