Generation and protective efficacy of a cold-adapted attenuated genotype 2b porcine epidemic diarrhea virus

Won, Hokeun; Lee, Dong-Uk; Jang, Guehwan; Noh, Yun-Hee; Lee, Seung-Chul; Choi, Hwan-Won; Yoon, In-Joong; Yoo, Han Sang; Lee, Changhee

The cold-adapted G2b PEDV Aram-CA strains were generated by a serial passage of Aram-P5 at low temperatures commencing from 37°C to 32°C and additional adaptation for growth up 45 passages at 32°C in Vero cells. We initially aimed to evaluate the immunogenicity of the cold-adapted Aram derivatives in the natural host. Antisera were collected from the pigs at 0-, 1-, 2-, and 3-week after oral administration of each virus and were tested for neutralizing activity against the Aram isolate. The serum samples obtained from pigs immunized with the cold-adapted Aram-P29-CA strain had higher neutralizing antibody titers, when compared with those from pigs in other CA virus-inoculated groups (Fig. 4) . Furthermore, the immune sera of pigs inoculated with Aram-P40-CA or -P50-CA strain showed little or no neutralizing activity against PEDV. Since our data indicated that only Aram-P29-CA can elicit robust antibody responses in immunized animals, this cold-adapted virus was selected for subsequent in vitro and in vivo studies. compared to 32°C, indicating impairment of growth at 37°C. This diminished growth at 37°C was confirmed by comparing growth kinetics in Vero cells at both the temperatures. The Aram-P29-CA strain exhibited efficient growth and had an infectious titer of > 10 6.5 TCID 50 / mL at 32°C by 2 dpi, but slower replication rates with a maximum titer of 10 2.5 TCID 50 /mL than that of the parental Aram-P5 strain (a 10,000-fold reduction) at 37°C, indicating an adaptation to low temperatures.