Publications

4498

Huanglongbing (HLB) is the most destructive, yet incurable disease of citrus. Finding sources of genetic resistance to HLB-associated ‘CandidatusLiberibacter asiaticus’ (Las) becomes strategic to warrant crop sustainability, but no resistantCitrusgenotypes exist. SomeCitrusrelatives of the family Rutaceae, subfamily Aurantioideae, were described as full-resistant to Las, but they are phylogenetically far, thus incompatible withCitrus. Partial resistance was indicated for certain cross-compatible types. Moreover, other genotypes from subtribe Citrinae, sexually incompatible but graft-compatible withCitrus, may provide new rootstocks able to restrict bacterial titer in the canopy. Use of seedlings from monoembryonic species and inconsistencies in previous reports likely due to Las recalcitrance encouraged us to evaluate more accurately theseCitrusrelatives. We tested for Las resistance a diverse collection of graft-compatible Citrinae species using an aggressive and consistent challenge-inoculation and evaluation procedure. Most Citrinae species examined were either susceptible or partially resistant to Las. However,Eremocitrus glaucaand Papua/New GuineaMicrocitrusspecies as well as their hybrids and those withCitrusarose here for the first time as full-resistant, opening the way for using these underutilized genotypes as Las resistance sources in breeding programs or attempting using them directly as possible new Las-resistantCitrusrootstocks or interstocks.

Abstract BACKGROUND: Rickettsia spp. are human pathogens that cause a number of diseases and are transmitted by arthropods, such as ixodid ticks. Estonia is one of few regions where the distribution area of two medically important tick species, Ixodes persulcatus and I. ricinus, overlaps. The presence of the nidicolous rodent-associated I. trianguliceps has also recently been shown in Estonia. Although there is no data available in Estonia on human disease caused by tick-borne Rickettsia spp., the presence of three Rickettsia species in non-nidicolous ticks was also previously reported. The aim of this study was to detect, identify and partially characterize Rickettsia species in nidicolous and non-nidicolous ticks attached to rodents.RESULTS: Larvae and nymphs of I. ricinus (n = 1004), I. persulcatus (n = 75) and I. trianguliceps (n = 117) removed from rodents and shrews caught in different parts of Estonia were studied for the presence of Rickettsia spp. by nested PCR. Ticks were collected from 314 small animals of 5 species (Myodes glareolus (bank voles), Apodemus flavicollis (yellow necked mice), A. agrarius (striped field mice), Microtus subterranius (pine voles) and Sorex araneus (common shrews)). Rickettsial DNA was detected in 8.7% (103/1186) of the studied ticks. In addition to R. helvetica previously found in questing ticks, this study reports the first identification of the recently described I. trianguliceps-associated Candidatus R. uralica west of the Ural Mountains.

HighlightsA portable system based on real-time fluorescence analysis was developed for field detection of Candidatus Liberibacter asiaticus within 40 min from sample to answer.A smartphone-assisted device was designed for easy operation, reliable nucleic acid amplification, and highly sensitive fluorescence detection, with sensitivity comparable to that of a commercial instrument.A novel homemade 3D printed box was used for in-field reagent storage, and it could maintain low temperature (<4°C) for about 8 hours without power supply.This fully integrated system is stable, easy to use, inexpensive, and has great application prospects in resource-limited areas.Abstract. Candidatus Liberibacter asiaticus (Las) is a main causal agent of huanglongbing (HLB), a destructive disease that has greatly reduced citrus yields and quality. Instruments with high sensitivity and portability are urgently required for on-site testing. In this study, a novel sample-to-answer optical system for on-site detection of Las was developed. Three major functions, including DNA extraction, amplification, and detection, are integrated into a portable case. This system mainly consists of (1) a specially designed 3D printed box for on-site reagent storage that can maintain low temperature (below 4°C) for 7.5 h at ambient temperature (35°C); (2) a custom device, called the IF-Device, for DNA amplification and detection of HLB, with an optimized optical structure, a sensitive signal processing circuit, and a precise temperature control algorithm with an accuracy of ±0.1°C; and (3) a battery-based power supply for the whole system. In a typical test using sodium fluorescein as a standard model, the results showed that the sensitivity of this system (1.0 nM) could easily meet the requirements of fluorescence biosensors. The feasibility of this homemade system was evaluated with samples extracted from infected citrus leaves based on the loop-mediated isothermal amplification (LAMP) method, and the limit of detection (LOD) was approximately 1.0 × 10-4 ng µL-1. The whole detection process for eight samples could be simultaneously accomplished within 40 min, and the results could be displayed on a smartphone in real-time. Moreover, the portable case is anti-interference, low cost, and only 2 kg in weight. Considering its sensitivity, stability, and portability, this highly integrated system possesses promising prospects for in-field detection. Keywords: Field detection, Fluorescence biosensor, Huanglongbing, Isothermal amplification, Sample-to-answer.

‘Candidatus Liberibacter asiaticus’ (CLas) is an unculturable phloem-restricted α-proteobacterium associated with huanglongbing (HLB). Here, we provide the genome sequence of CLas strain CoFLP1 from its insect vector Diaphorina citri (Hemiptera: Liviidae) collected in the department of La Guajira, Colombia. The CoFLP1 strain is composed of 1,231,639 bp with G+C 36.5% content. This study reports the first CLas genome sequence from Colombia, which will add to CLas genome resources and help to elucidate our understanding of the introduction pathway of HLB in South America.

iaphorina citri is considered the most dangerous citrus pest because it transmits Candidatus Liberibacter asiaticus, the causal agent of Huanglongbing (HLB). Like other hemiptera insects, D. citri has developed mutualistic interactions with prokaryotic organisms known as endosymbionts. This symbiosis can be obligatory, when the interaction occurs with a primary endosymbiont, or facultative, when it is a secondary endosymbiont. Symbiosis is essential for various physiological functions, but some endosymbionts can adversely affect the psyllid's abilities. D. citri is associated with a great diversity of endosymbionts, with Candidatus Carsonella ruddii, Candidatus Profftella armatura, Candidatus Wolbachia spp. and Candidatus Liberibacter spp standing out. The aim o symbiotic relationship, prokaryotes, Candidatus Wolbachia, transmission of Candidatus Liberibacter asiaticus.