Chikungunya (CHIK) is an emerging or reemerging arboviral infectious disease transmitted to humans by the Asian tiger mosquito, Aedes albopictus (Skuse), and the yellow fever mosquito, Ae. aegypti (L.), particularly in Africa, India, Southeast Asia, and other parts of Europe where numerous outbreaks and epidemics were documented during recent decades. This viral disease is a potential threat to human health worldwide; thus, a quantitative one-step, real-time RT-PCR protocol was developed and assessed as a suitable assay for minimum detection of chikungunya virus (CHIKV) E1 (genus Alphavirus, family Togaviridae) in human serum (7.51 fg/μl) by using the QuantiTect^® Probe RT-PCR kit with appropriate synthetic primer (CHIK E1 F and CHIK E1 R) and dual-labeled CHIK E1 probe. Unfortunately, we did not detect any CHIKV from 9 species, namely Ae. albopictus, Aedes (=Ochlerotatus) cantans (Meigen), Ae. cinereus Meigen, Ae. geminus Peus, Ae. vexans (Meigen), Anopheles claviger Meigen, An. maculipennis Meigen s.l., An. plumbeus Stephens, and Culex pipiens L., but some Kamiti River virus (KRV) were found in Ae. albopictus eggs, Ae. cinereus and Ae. vexans mosquitoes adults. Phylogenetic analyses revealed that these novel insect flaviviruses from Ae. albopictus eggs is closely related to Cell fusing agent virus (CFAV), which is different from the other group of arboviruses from extra Aedes spp. mosquitoes. These results suggest that Aedes spp. in the area of Ticino, Switzerland are likely to be highly infected with two distinct groups of insect flaviviruses, excluding the possibility of CHIKV in mosquito vectors.