Publications

3846

Huanglongbing (HLB) is a difficult-to-control and highly destructive citrus disease that, in Brazil, is associated mainly with the bacterium ‘Candidatus Liberibacter asiaticus’ transmitted by the psyllid Diaphorina citri. The aim of this study was to improve our understanding of the ‘Ca. L. asiaticus’ infection process by exposing excised, fully expanded, immature citrus leaves in 50-ml Falcon tubes to one, four, or eight adults from a ‘Ca. L. asiaticus’-exposed colony for 1-, 3-, 7-, or 15-day periods for access to inoculation (IAP). The leaves were incubated at 26°C for 1, 3, 7, 15, and 21 days (incubation period [IP]). Infection frequencies and ‘Ca. L. asiaticus’ titers were assessed by quantitative polymerase chain reaction (qPCR). ‘Ca. L. asiaticus’ infection was a function of leaf age, number of insects, IAP, and IP. In general, higher infection rates were observed on younger leaves inoculated with higher numbers of insects and after longer IAP and IP. The immature excised leaf method allowed determination of 3 to 7 days as the range of time required by ‘Ca. L. asiaticus’ to reach qPCR detectable levels. Even though leaf survival could be prolonged by the maintenance of a branch segment at the base of the leaf petiole, leaf degradation, visible after about 15 days IP, did not allow observation of the entire infection process which, in the intact plant, culminates with the appearance of the blotch mottling symptom on leaf blades.

Although ‘Candidatus Liberibacter asiaticus’ (Las) is a major pathogen associated with citrus huanglongbing (HLB), some characteristics of transmission by the psyllid vector Diaphorina citri are not fully understood. We examined the latent period and persistence of transmission of Las by D. citri in a series of experiments at 25°C, in which third-instar psyllid nymphs and 1-week-old adults were confined on infected citrus for an acquisition access period (AAP), and submitted to sequential inoculation access periods (IAPs) on healthy citrus seedlings. The median latent period (LP50, i.e., acquisition time after which 50% of the individuals can inoculate) of 16.8 and 17.8 days for psyllids that acquired Las as nymphs and adults, respectively, was determined by transferring single individuals in 48-h IAPs. Inoculation events were intermittent and randomly distributed over the IAPs, but were more frequent after acquisition by nymphs. A minimum latent period of 7 to 10 days was observed by transferring groups of 10 psyllids in 48-h IAPs, after a 96-h AAP by nymphs. Psyllids transmitted for up to 5 weeks, when submitted to sequential 1-week IAPs after a 14-day AAP as nymphs. The long latent period and persistence of transmission are indirect evidences of circulative propagation of Las in D. citri.

Significance A wide range of organisms sense Earth’s magnetic field for navigation. For some organisms, like magnetotactic bacteria, magnetic particles form inside cells and act like a compass. However, the origin of magnetotactic behavior remains a mystery. We report that magnetotaxis evolved in bacteria during the Archean, before or near the divergence between the Nitrospirae and Proteobacteria phyla, suggesting that magnetotactic bacteria are one of the earliest magnetic-sensing and biomineralizing organisms on Earth. The early origin for magnetotaxis would have provided evolutionary advantages in coping with environmental challenges faced by microorganisms on early Earth. The persistence of magnetotaxis in separate lineages implies the temporal continuity of geomagnetic field, and this biological evidence provides a constraint on the evolution of the geodynamo.

Huanglongbing also known in Indonesia as citrus vein phloem degeneration (CVPD) is a devastating disease in citrus plantation worldwide, especially in Asia, Africa, and America. In Asian countries including Indonesia, Candidatus Liberibacter asiaticus (CLas) has been confirmed as the causal agent of huanglongbing disease on citrus. Distribution of CLas in Indonesia has been reported in West Borneo, East Nusa Tenggara, Bali, Yogyakarta, Central Java and East Java.  The purpose of this study was to detect CLas in several Indonesia’s citrus plantations that has not and has been reported previously and to study its genetic diversity and their relationship. DNA of plant samples, i.e. citrus leaves, was extracted using CTAB method and CLas was amplified using PCR with the A2/J5 primer pair. DNA amplification results showed that infection of CLas was positively detected from samples from Bogor and Cibodas (West Java), Tuban and Jember (East Java), as well as Katung, Bayung Gede, Kerta, and Pancasari (Bali). Alignment of nucleotide sequences from positive samples showed that their ribosomal protein β-operon has high similiarity to that of CLas OK901 originated from Okinawa (Japan). Isolates of CLas originated from Bogor, Cibodas, Tuban, Jember, and Katung have been known to be identical to other CLas of Indonesian origins reported earlier.  Isolates of CLas originated from Bayung Gede, Pancasari, and Kerta have single nucleotide polymorphisms at 6 points of bases of the 539 total bases compared in their conservative regions, although only 3 of the 6 bases could affect their amino acid sequences.