Putative Novel Viruses in the Families Lispiviridae and Rhabdoviridae Detected in Culex and Anopheles Mosquitoes Collected at the São Paulo Zoo

Several viruses belonging to the order Mononegavirales are recognized as highly pathogenic in humans and are often associated with lethal disease outcomes. Prominent examples include human respiratory syncytial virus (HRSV), Ebola virus (EBOV), rabies virus (RABV), and Marburg virus (MARV). In addition to their clinical severity, these viruses are considered potential biological threats due to their high transmissibility and virulence. In this study, we performed a metatranscriptomic analysis of Culex and Anopheles mosquitoes collected at the DBB (Diretoria de Biodiversidade e Biotecnologia), São Paulo, Brazil. Our analysis revealed viral sequences associated with two families within Mononegavirales : Lispiviridae and Rhabdoviridae . Phylogenetic analysis of the Lispiviridae sequences identified six variants, designated CxLispV-SP_03 , CxLispV-SP_09 , CxLispV-SP_12 , CxLispV-SP_13 , CxLispV-SP_14 , and CxLispV-SP_15 , that formed a strongly supported monophyletic clade with Canya virus, suggesting the discovery of a putative novel viral lineage. Examination of RNA‐dependent RNA polymerase (RdRp) domains in these sequences confirmed the presence of essential catalytic motifs. In contrast, the detected rhabdoviruses exhibited greater genomic structural heterogeneity, including the presence of accessory protein domains. Pairwise comparisons based on RdRp amino acid identity delineated four distinct groups among these sequences, pointing to substantial evolutionary divergence within the sampled rhabdovirus diversity. Together, these findings contribute to the growing characterization of mosquito‐associated mononegaviruses and provide a genomic foundation to support future surveillance efforts and public health risk assessments related to vector‐borne viruses.