Molecular mechanisms underlying heat or tetracycline treatments for citrus HLB control

Authors: DING, FANG - US Department Of Agriculture (USDA); Allen, Victoria; ZHANG, SHOUAN - University Of Florida; Duan, Ping; LOU, WEIQI - North Carolina State University

Submitted to: Horticulture Research
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 3/27/2018
Publication Date: 6/1/2018
Citation: Ding, F., Allen, V.W., Zhang, S., Duan, Y., Lou, W. 2018. Molecular mechanisms underlying heat or tetracycline treatments for citrus HLB control. Horticulture Research. 5:30. https://doi.org/10.1038/s41438-018-0038-x.
DOI: https://doi.org/10.1038/s41438-018-0038-x

Interpretive Summary: Citrus Huanglongbing (HLB), also known as citrus greening, is one of the most destructive diseases that affects the citrus industry throughout the world. Our previous study showed that controlled heat treatment dramatically reduced Las titer in both citrus and periwinkle plants infected with HLB bacterium, with Las becoming undetectable months after heat treatment. A follow up proteomics study demonstrated that heat treatment causes several chaperone-related proteins to be up-regulated in citrus trees. More recently, we analyzed the transcriptome of heat treated trees and showed that in HLB-affected citrus, post-treatment gene expression more closely resembled that of healthy controls. In this study, we investigated the molecular mechanism underlying heat or tetracycline treatments on the HLB bacterium, ‘Candidatus Liberibacter asiaticus’ (Las) by focusing on Las prophage/phage conversion under stress conditions. By comparing the prophage FP1 and FP2 copy number to the copy number of 16S rDNA in HLB-affected plants, we found the relative copy number of both FP1 and FP2 increased significantly when the temperature was increased from 23°C to 37°C, 42°C or 45°C. When treated with tetracycline at 50 - 150 µg/ml and 200 - 250 µg/ml, respectively, the relative copy number of both FP1 and FP2 increased by 3.4 to 6.0 fold. Furthermore, using transmission electron microscopy, we observed increased number of phage particles upon temperature increase , providing direct evidence of lysogenic to lytic conversion. These results not only provide new insight into the molecular mechanisms underlying heat or tetracycline treatment, but also suggest a novel HLB control strategy by enhancing the endogenous conversion from Las prophages to phages.

Technical Abstract: Huanglongbing (HLB), a destructive bacterial disease, severely impedes worldwide citrus production. In our previous reports, we revealed the molecular mechanisms of host plant responses that underlie thermotherapy against HLB. In this study, we investigated the molecular mechanism underlying heat or tetracycline treatments on the HLB bacterium, ‘Candidatus Liberibacter asiaticus’ (Las) by focusing on Las prophage/phage conversion. By comparing the prophage FP1 and FP2 copy number to the copy number of 16S rDNA in HLB-affected plants, we found the relative copy number of both FP1 and FP2 increased significantly, ranging from 3.4 to 6.7 fold change when the temperature was increased from 23°C to 37°C, 42°C or 45°C. When treated with tetracycline at 50 - 150 µg/ml and 200 - 250 µg/ml, respectively, the relative copy number of both FP1 and FP2 increased by 3.4 to 6.0 fold. Meanwhile, Las prophage structural gene and antirepressor gene copy numbers showed similar trends. Furthermore, transmission electron microscopy provided direct evidence of lysogenic to lytic conversion upon temperature increase. These results not only provide new insight into the molecular mechanisms underlying heat or tetracycline treatment, but also suggest a novel HLB control strategy by enhancing the endogenous conversion from Las prophages to phages.