Citrus yellow vein clearing virus （CYVCV） is considered as the causal agent of Citrus yellow vein clearing disease and belongs to the genus Mandarivirus in the family Alphaflexiviridae. Capsid protein （CP） of CYVCV Chongqing isolate （CYVCV- CQ） was produced using a prokaryotic expression system and used as the immunogen for monoclonal antibody （MAb） production. Four highly specific and sensitive murine MAbs and one polyclonal antibody were prepared in this study. Titers of the four MAbs in ascites fluids ranged from 10-6 to 10-7 as determined by indirect enzyme-linked immunosorbent assay （ELISA）. Three serological assays, including dot enzyme-linked immunosorbent assay （dot-ELISA）, tissue blot-ELISA, and double-antibody sandwich （DAS）-ELISA, were developed for quick and reliable detections of CYVCV in citrus samples. The developed dot-ELISA and DAS-ELISA methods could detect CYVCV in the infected citrus leaf crude extracts diluted at 1：2 560 and 1：10 240 （w/v, g mL^-1）, respectively. The detection result of 125 citrus leaf samples collected from citrus groves in Yunnan Province and Chongqing Municipality of China showed that approximately 36% samples were positive for CYVCV. This virus was, however, not＇detected in any sample collected from Zhejiang or Jiangxi Province, China.
LIU ZhenSUNZHU Yuan-jiZHOU Xue-pingHONG JianWU Jian-xiang
Plant intracellular nucleotide binding leucine-rich repeat (NLR) immune receptors play critical roles in pathoge n surveillance. Most plant NLRs characterized so far were found to use a single domain/sensor to recognize pathogen effectors. Here we report that the Sw-5b NLR immune receptor uses two distinct domains to detect the viral movement protein NSm encoded by tospovirus. In addition to its leucine-rich repeat (LRR) domain that has been previously reported, the N-terminal Solanaceae domain (SD) of Sw- 5b also interacts with NSm and a conserved 21-amino-acid region of NSm (NSm^21). The specific interaction between Sw-5b SD and NSm is required for releasing the inhibitory effect of coiled-coil domain on the NBARC- LRR region. Furthermore, we found that the binding of NSm affects the nucleotide binding activity of the NB-ARC-LRR in vitro, while Sw-5b NB-ARC-LRR is activated only when NSm and NSm^21 levels are high. Interestingly, Sw-5b SD could significantly enhanee the ability of the NB-ARC-LRR to detect low levels of NSm effector and facilitate its activation and induction of defense response. An Sw-5b SD mutant that is disrupted in NSm recognition failed to enhance the ability of the NB-ARC-LRR to sense low levels of NSm and NSm^21 . Taken together, our results suggest that Sw-5b SD functions as an extra sensor and the NB-ARC-LRR as an activator, and that Sw-5b NLR adopts a two-step recog nition mechanism to enhance viral effector perception.
Jia LiHaining HuangMin ZhuShen HuangWenhua ZhangSavithramma P. Dinesh-KumarXiaorong Tao
