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Huanglongbing (HLB) is a globally devastating disease of citrus. Presently, three etiological agents are associated with HLB and include; Candidatus Liberibacter asiaticus (CLas), Candidatus Liberibacter americanus; and Candidatus Liberibacter africanus. Attempts to determine alternate (non-Liberibacter) associated etiological agents of HLB have been performed, namely by metagenomic analyses with HLB phenotypic citrus of phloem tissue isolated from bark [1] and whole leaf midribs [2]. These reports indicated a strong correlation for Liberibacter species associated with HLB etiology, but they do not indicate the presence of other significant associated etiological agents. Utilizing both PCR and non-PCR based metagenomic strategies; these previous reports present an undersized view (relating to inherent technique limits, restricted sample scope, and/or bacterial bias) into the etiology of HLB. Here we report ancillary contiguous metagenomic sequences contained amongst whole CLas genome amplification contiguous sequences that were sequenced on the next generation Ion Torrent PGM sequencing system. Within these ancillary sequences, a diverse metagenomic community is present, indicating a rich diversity of both prokaryotic and eukaryotic organisms that are unique and in common within these globally isolated HLB citrus samples. Although this report does not represent a complete metagenomic study of HLB diseased citrus, it does suggest that the use of Ion Torrent PGM sequencing system can be employed for metagenomic analysis. These findings justify a more complete analysis of the metagenome of HLB etiology, which may help further elucidate the HLB disease complex.

Author(s): Bove, Josy | Abstract: It will be recalled how the notions of “heat-sensitive/heat-tolerant HLB” and “African HLB/Asian HLB” were developed. These notions benefited from the possibility to confirm, for the first time, the non-specific HLB symptoms by reliable laboratory techniques: detection of the HLB-associated bacterium by transmission electron microscopy from 1970 onwards to DNA-hybridization and PCR of the liberibacters in the 1990s. With these tools, the early history of citrus HLB in the many countries surveyed could be more precisely described. This presentation also shows or proposes why: (i) African HLB is heat-sensitive and Asian HLB, heat-tolerant; (ii) only one of the seven known liberibacters, namely Candidatus Liberibacter asiaticus (Las), is heat-tolerant, Candidatus Liberibacter africanus (Laf) being a heat-sensitive liberibacter; (iii) African HLB is native to Africa, and Asian HLB is native to Asia; (iv)n “Continental Drift” supports astonishingly well the presence of Laf in Africa andn that of Las in Asia; presence of Las in the Americas is the result of incursions; (v) HLBn is not native to the Arabian Peninsula, but is the result of African and Asian HLB incursions into the peninsula; vi) recent presence of Las in Ethiopia is also the result of an incursion; vii) Candidatus Liberibacter americanus (Lam) in South America is the result of an incursion too; viii) while Laf and Las are of Gondwanan origin, Candidatus Liberibacter europaeus