Temporal and spatial detection of Candidatus Liberibacter asiaticus putative effector transcripts during interaction with Huanglongbing-susceptible, -tolerant, and -resistant citrus hosts

Authors: Shi, Qingchun; PITINO, MARCO - University Of Florida; Zhang, Shujian; Krystel, Joseph; CANO, LILIANA - University Of Florida; Shatters, Robert - Bob; Hall, David; Stover, Eddie

Submitted to: BMC Plant Biology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 3/6/2019
Publication Date: 4/3/2019
Citation: Shi, Q., Pitino, M., Zhang, S., Krystel, J.A., Cano, L.M., Shatters, R.G., Hall, D.G., Stover, E.W. 2019. Temporal and spatial detection of Candidatus Liberibacter asiaticus putative effector transcripts during interaction with Huanglongbing-susceptible, -tolerant, and -resistant citrus hosts. Biomed Central (BMC) Plant Biology. 19:122. https://doi.org/10.1186/s12870-019-1703-4.
DOI: https://doi.org/10.1186/s12870-019-1703-4

Interpretive Summary: Citrus Huanglongbing (HLB) is a devastating disease for the United States citrus industry. It is not clear how the pathogen/citrus interaction results in pathogen establishment and difficult to research because this pathogen cannot be grown outside of the hosts. Here we analyzed a group of possible disease-causing genes called effectors in the pathogen for their expression in the infected citrus plants. Types of citrus (susceptible, tolerant and resistant) and tissue (root vs. leaves) significantly varied the expression levels of some of the effector genes. Highly expressed effectors are more likely to be associated with HLB disease and merit further study.

Technical Abstract: Citrus Huanglongbing (HLB) is a bacterial disease with high economic significance. The associated agent Candidatus Liberibacter asiaticus (Ca. L.) is a fastidious, phloem-limited, intracellular bacterium that is transmitted by an insect vector the Asian citrus psyllid (ACP). The genome of Ca. L. asiaticus contains protein secretion machinery that suggests host cell modulation capacity of this bacterium. In this study, a total of 28 candidate effectors, an important class of secreted proteins, were predicted from the Ca. L. asiaticus genome. Sequence specific primers were designed for reverse transcription (RT) and quantitative polymerase chain reaction (qPCR), and expression was validated for 20 of the effector candidates in infected citrus with multiple genetic background. Using detached leaf inoculation, the messenger Ribonucleic acid interference (mRNA) of effectors was detected from 6 hours to 7 days post ACP exposure. It was observed that higher bacterial titers were associated with a larger number of effectors showing amplification across all samples. The effectors’ expression were compared in citrus hosts with various levels of HLB tolerance, including susceptible Duncan grapefruit and Washington navel orange, tolerant citron and Cleopatra mandarin, and resistant Pomeroy trifoliate and Carrizo citrange. Across all genotypes relatively high expression was observed for CLIBASIA_03695, CLIBASIA_00460, CLIBASIA_00420, CLIBASIA_04580, CLIBASIA_05320, CLIBASIA_04425, CLIBASIA_00525 and CLIBASIA_05315 in either a host-specific or -nonspecific manners. The two genotypes in each HLB-response group also show effector-expression profiles that seem to be different. In a companion study, the expression of effectors was compared between leaves and roots of own-rooted citrus that had been Ca. L. asiaticus-infected for more than a year. Results indicated relatively high expression of CLIBASIA_03875, CLIBASIA_04800 and CLIBASIA_05640 in all leaf and some root tissues of citron, Duncan and Cleopatra. In summary, this temporal and spatial expression analysis of Ca. L. asiaticus effectors identified candidates possibly critical for early bacterial colonization, host tolerance suppression and long-term survival which are all worthy of further investigation.