Objective To identify the blood meal source in Culicoides using polymerase chain reaction (PCR), and to study the diversity of blood-sucking habit. Methods Blood-engorged Culicoides were collected by light-traps in habitats related to human activities at a village in suburban Nanchang, China, August 2017. Species-specific primers were designed based on differences between mitochondrial cytochrome b sequences of common Culicoides hosts to set up a PCR system. The blood meal source in Culicoides was identified with positive and negative controls of animal blood source. Results Culicoides arakawae and C. oxystoma were the predominant species in the peasant household habitat, accounting for 37.01% and 33.85%, respectively. Culicoides arakawae (81.40%) was the dominating specie in chicken coops and C. oxystoma (57.66%) was the predominant specie in pigsties. Results of PCR for blood meal source in Culicoides showed that specific PCR products were amplified from the DNA extracted from Culicoides and known animal blood samples. Moreover, C. arakawae, C. oxystoma, C. punctatus, and C. homotomus could suck both human and animal blood, and they had the habit of simultaneously sucking blood from 2-4 host animals. The blood sucking index for human blood of C. arakawae, C. oxystoma, C. punctatus, and C. homotomus was 0.13, 0.63, 0.33, and 1.00, respectively. Conclusion The PCR method has advantages in sensitivity and reliability in identifying the Culicoides blood meal source. The four Culicoides are all capable of feeding on human and animal blood, with wild blood-sucking objects and diverse habit.