| Psittacine beak and feather disease(PBFD)is a contagious disease characterized by feather degeneration and malnutrition as well as beak deformity.This disease is caused by the psittacine beak and feather disease virus(PBFDV),which belongs to circovirus family.It is an icosahedral,symmetric,single-stranded and non-enveloped DNA virus with a genome of approximately 2 kb.PBFDV can infect multiple types of parrots at all age stages.Moreover,it can also infect other species of birds.Infected adult birds usually show no signs of clinical symptoms due to the mature immune system.However,infected juvenile birds generally exhibit signs of clinical symptoms,such as feather loss and beak deformity.Since the virus could lead to parrot immune-suppression and damage the immune system,numerous infected parrots finally die from secondary infections.In the middle of March 2017,a psittacine beak and feather disease epidemic struck a farm in Fuzhou.Dozens of parrots(including honey-sucking parrots,baby sun parrots and macaws)died one after another with some typical symptoms.These cases were comprehensively diagnosed and analyzed by pathological anatomy,histopathological observation and PCR examination.Through anatomy,multiple organs were found to exhibit signs of pathological changes of varying degrees.Further analysis of histopathology displayed liver necrosis as well as edema;intestinal inflammation;tubular epithelial cells shedding and necrosis;spleen necrosis,edema and so on.In order to determine the molecular characteristics of this strain,the whole-genome sequences were obtained by PCR amplification followed by DNA sequencing,and this PBFDV was provisionally named as FZ strain.Moreover,a phylogenetic tree according to the whole-genome demonstrated that FZ strain was closely related to and located in the same branch with these strains from New Caledonia rather than those previously reported from Qingdao in China.Meanwhile,by comparing FZ strain with other strains,the nucleotide homology ranged from 83.6%to 93.22%.In addition,the genes encoding capsid protein(Cap)and replication-associated protein(Rep)were successfully cloned into the Escherichia coli expression vector to reconstruct pGEX-4T-1-Cap and pGEX-4T-1-Rep based on the whole-genome sequence of the strain.Induced by IPTG,GST fusion proteins(GST-Cap and GST-Rep)of 54 kDa and 59 kDa were successfully expressed in Escherichia coli BL21.Furthermore,polyclonal antibodies were successfully generated by immunizing rabbits with GST-Cap fusion protein.Using western blotting to detect the dead parrot tissues infected by the PBFDV,a positive band was observed at a size of 28 kDa.These results not only indicated that the antibodies had good reaction specificity,but also confirmed that the above cases were indeed infected with the PBFDV.In conclusion,we first isolated and identified a PBFDV strain in Fujian Province that is closely related to PBFDV strains from New Caledonia.We have observed that this PBFDV strain caused pathological changes of various organs of the infected parrots.These results suggest that this strain may have high pathogenicity in parrots.In addition,the anti-PBFDV Cap antibodies we prepared had good specificity and could be further used to develop diagnostic kits for clinical detection and further studies of PBFDV pathogenesis and its protein Cap. |
