The environmental conditions of chicken houses play an important role in the growth and development of these animals. The chicken house is an essential place for the formation of microbial aerosols. Microbial aerosol pollution and transmission can affect human and animal health. In this work, we continuously monitored fine particulate matter (PM2.5) in the chicken house environment for four weeks and studied the microbial community structure in the aerosols of the chicken house environment through metagenomic sequencing. Our results found that bacteria, fungi, viruses, and archaea were the main components of PM2.5 in the chicken house environment, accounting for 89.80%, 1.08%, 2.06%, and 0.49%, respectively. Conditional pathogens are a type of bacteria that poses significant harm to animals themselves and to farm workers. We screened ten common conditional pathogens and found that Staphylococcus had the highest relative abundance, while Clostridium contained the most microbial species, up to 456. Basidiomycetes and Ascomycota in fungi showed dramatic changes in relative abundance, and other indexes showed no significant difference. Virulence factors (VF) are also a class of molecules produced by pathogenic microbes that can cause host diseases. The top five virulence factors were found in four groups: FbpABC, HitABC, colibactin, acinetobactin, and capsule, many of which are used for the iron uptake system. In the PM2.5 samples, eight avian viruses were the most significant discoveries, namely Fowl aviadovirus E, Fowl aviadovirus D, Avian leukosis virus, Avian endogenous retrovirus EAV-HP, Avian dependent parvovirus 1, Fowl adenovus, Fowl aviadovirus B, and Avian sarcoma virus. The above results significantly improve our understanding of the microbial composition of PM2.5 in chicken houses, filling a gap on virus composition; they also indicate a potential threat to poultry and to human health. This work provides an important theoretical basis for animal house environmental monitoring and protection.