Publications

4478

‘Candidatus Liberibacter solanacearum’ is a plant pathogen affecting the families Solanaceae and Apiaceae in different parts of the world. ‘Ca. L. solanacearum’ is a Gram-negative, fastidious α-proteobacterium that is vectored by different psyllid species. Plant-pathogenic bacteria are known for interfering with the host physiology or defense mechanisms, often by secreting bacterial effectors. Effector proteins are critical for virulence; therefore, the identification of effectors could help with disease management. In this study, we characterized the Sec-translocon-dependent ‘Ca. L. solanacearum’–hypothetical protein effector 1 (Lso-HPE1). We compared this protein sequence in the different ‘Ca. L. solanacearum’ haplotypes. We predicted the signal peptide and validated its function using Escherichia coli’s alkaline phosphatase fusion assay. Agrobacterium tumefaciens-mediated transient expression in Nicotiana benthamiana demonstrated that Lso-HPE1 from ‘Ca. L. solanacearum’ haplotypes A and B were able to inhibit the induction of cell death in plants. We also compared gene expression of the Lso-HPE1- transcripts in ‘Ca. L. solanacearum’ haplotypes A and B in tomato and in the vector Bactericera cockerelli. This work validates the identification of a Sec-translocon-dependent ‘Ca. L. solanacearum’ protein possibly involved in suppression of plant cell death.

‘Candidatus Liberibacter asiaticus’ (Las) is an unculturable α-proteobacterium associated with citrus huanglongbing (HLB), a devastating disease currently threatening the citrus industry worldwide. Here, we present the genome sequence of Las strain TaiYZ2 from an HLB-affected pomelo tree in Hat Yai district, Songkhla Province, Thailand. The TaiYZ2 genome is composed of 1,230,623 bp with G+C content of 36.4%. This is the first Las genome sequence from Thailand, which will enrich current Las genome resource and facilitate HLB research and management.

‘Candidatus Liberibacter asiaticus’ is the unculturable causative agent of citrus huanglongbing disease. Here, we report the first citrus root metagenome sequence containing the draft genome of ‘Ca. L. asiaticus’ strain AHCA17, obtained from a pummelo tree in California. The assembled genome was 1.2 Mbp and resulted in 37 contigs (N50 = 158.7 kbp) containing 1,057 predicted open reading frames and 45 RNA-coding genes. This draft genome will provide a valuable resource in further study of ‘Ca. L. asiaticus’ genome diversity and pathogen epidemiology.

A novel actinobacterial strain, designated K81G1T, was isolated from a soil sample collected in Kantulee peat swamp forest, Surat Thani Province, Thailand, and its taxonomic position was determined using a polyphasic approach. Optimal growth of strain K81G1Toccurred at 28–30 °C, at pH 5.0–6.0 and without NaCl. Strain K81G1Thad cell-wall chemotype IV (meso-diaminopimelic acid as the diagnostic diamino acid, and arabinose and galactose as diagnostic sugars) and phospholipid pattern type II, characteristic of the genusAmycolatopsis. It contained MK-9(H4) as the predominant menaquinone, iso-C16 : 0, C17 : 0cyclo and C16 : 0as the major cellular fatty acids, and phospholipids consisting of phosphatidylglycerol, diphosphatidylglycerol, phosphatidylethanolamine, hydroxyphosphatidylethanolamine, phosphatidylinositol and two unidentified phospholipids. Based on 16S rRNA gene sequence similarity and phylogenetic analyses, strain K81G1Twas most closely related toAmycolatopsis rhizosphaeraeTBRC 6029T(97.8 % similarity),Amycolatopsis acidiphilaJCM 30562T(97.8 %) andAmycolatopsis bartoniaeDSM 45807T(97.6 %). Strain K81G1Texhibited low average nucleotide identity and digital DNA–DNA hybridization values withA. rhizosphaeraeTBRC 6029T(76.4 %, 23.0 %),A. acidiphilaJCM 30562T(77.9 %, 24.6 %) andA. bartoniaeDSM 45807T(77.8 %, 24.3 %). The DNA G+C content of strain K81G1Twas 69.7 mol%. Based on data from this polyphasic study, strain K81G1Trepresents a novel species of the genusAmycolatopsis, for which the nameAmycolatopsis acidicolasp. nov. is proposed. The type strain is K81G1T(=TBRC 10047T=NBRC 113896T).

Abstract The potato/tomato psyllid Bactericera cockerelli (Šulc) transmits ‘Candidatus Liberibacter solanacearum’ (Lso) (also known as ‘Candidatus Liberibacter psyllaurous’), the bacterium associated with zebra chip disease (ZC) in potato. When disease incidence is high, ZC causes large economic losses through reductions in potato yield and tuber quality. No commercial potato variety has been found totally resistant to the pathogen. We evaluated host acceptance behaviors using no-choice assays on three breeding clones derived from Solanum chacoense Bitter with putative tolerance to Lso and/or ZC as part of an effort to determine whether the disease tolerance observed in those breeding clones was related to effects on psyllid settling behavior. We also counted the number of eggs laid and nymphs hatched on the different genotypes to observe any differences in reproduction. The potato variety ‘Russet Burbank’ was used as a susceptible control. Probing frequency and female walking duration were greater on Russet Burbank than the other genotypes. Oviposition did not differ among genotypes. However, female psyllids on two of the Lso-tolerant genotypes displayed reduced fertility 18–24 d after confinement with a male, relative to females on Russet Burbank. These results suggest that although the germplasms display minor abiotic activity on psyllid fertility, tolerance to Lso may be more strongly linked with plant tolerance to the pathogen rather than effects on host acceptance behaviors.

Citrus huanglongbing (HLB), caused by phloem-limited ‘Candidatus Liberibacter’ bacteria, is a destructive disease threatening the worldwide citrus industry. The mechanisms of pathogenesis are poorly understood and no efficient strategy is available to control HLB. Here, we used a comparative genomics screen to identify candidate microbe-associated molecular patterns (MAMPs) from ‘Ca. Liberibacter’ spp. We identified the core genome from multiple ‘Ca. Liberibacter’ pathogens, and searched for core genes with signatures of positive selection. We hypothesized that genes encoding putative MAMPs would evolve to reduce recognition by the plant immune system, while retaining their essential functions. To efficiently screen candidate MAMP peptides, we established a high-throughput microtiter plate-based screening assay, particularly for citrus, that measured reactive oxygen species (ROS) production, which is a common immune response in plants. We found that two peptides could elicit ROS production in Arabidopsis and Nicotiana benthamiana. One of these peptides elicited ROS production and defense gene expression in HLB-tolerant citrus genotypes, and induced MAMP-triggered immunity against the bacterial pathogen Pseudomonas syringae. Our findings identify MAMPs that boost immunity in citrus and could help prevent or reduce HLB infection.