A Pathogen Secreted Protein as a Detection Marker for Citrus Huanglongbing

Abstract

The citrus industry is facing an unprecedented crisis due to Huanglongbing (HLB, aka citrus greening disease), a bacterial disease associated with the pathogen Candidatus Liberibacter asiaticus (CLas) that affects all commercial varieties. Transmitted by the Asian citrus psyllid (ACP), CLas colonizes citrus phloem, leading to reduced yield and fruit quality, and eventually tree decline and death. Since adequate curative measures are not available, a key step in HLB management is to restrict the spread of the disease by identifying infected trees and removing them in a timely manner. However, uneven distribution of CLas cells in infected trees and the long latency for disease symptom development makes sampling of trees for CLas detection challenging. Here, we report that a CLas secreted protein can be used as a biomarker for detecting HLB infected citrus. Proteins secreted from CLas cells can presumably move along the phloem, beyond the site of ACP inoculation and CLas colonized plant cells, thereby increasing the chance of detecting infected trees. We generated a polyclonal antibody that effectively binds to the secreted protein and developed serological assays that can successfully detect CLas infection. This work demonstrates that antibody-based diagnosis using a CLas secreted protein as the detection marker for infected trees offers a high-throughput and economic approach that complements the approved quantitative polymerase chain reaction-based methods to enhance HLB management programs.

Genome‐wide analyses of Liberibacter species provides insights into evolution, phylogenetic relationships, and virulence factors

“…Phytoplasma SDEs are able to move through plasmodesmata and the expression of individual SDEs in Arabidopsis thaliana mimics disease symptoms (Hoshi et al , ; MacLean et al , ). SDEs of Las have been explored to understand virulence, survival, and detection (Prasad et al , ; Pagliaccia et al , ; Shi et al , ). Las SDE1 targets citrus papain‐like cysteine proteases, which are induced during infection and known to be plant immune regulators (Clark et al , ).…”

“…Unlike most gram‐negative bacteria, Las lacks the type III secretion system, but possesses the general Sec secretion system, which is capable of secreting effectors directly outside bacterial cells (Sugio et al , ). Sec translocon‐dependent effectors (SDEs) are among the most likely Las virulence components (Pagliaccia, et al , ; Clark et al , ). However, the host targets of Las SDEs remain relatively unexplored.…”

“…However, the host targets of Las SDEs remain relatively unexplored. One Las SDE, SDE1 (CLIBASIA_05315), is primarily expressed in citrus and acts to inhibit defence‐induced host papain‐like cysteine protease activity (Pagliaccia, et al , ; Clark et al , ).…”

Transient expression of 'Candidatus Liberibacter asiaticus' effector SDE1 induces chlorosis by suppressing NbDDX3 gene in Nicotiana benthamiana

“…Sec-delivered effector 1 (SDE1) is a protein composed of 154 amino acids, and it can be secreted outside the cell through the Sec pathway upon expression in E. coli [32]. An SDE1-based polyclonal antibody has been successfully generated to diagnose and detect the bacterium owing to its speci city for Las pathogens [29]. The mature SDE1 protein without a signal peptide induces cell death in N. benthamiana in Agrobacterium-mediated transient transformation.…”

“…The mature SDE1 protein without a signal peptide induces cell death in N. benthamiana in Agrobacterium-mediated transient transformation. In contrast, the full length SDE1 lacks the ability to induce cell death [29]. In addition, the deletion of a chloroplast targeting sequence resulted in starch accumulation and chlorosis but not cell death [30].…”