scholarly journals Metabolome and Microbiome Signatures in the Roots of Citrus Affected by Huanglongbing

2019 ◽  
Vol 109 (12) ◽  
pp. 2022-2032 ◽  
Author(s):  
Emily M. T. Padhi ◽  
Nilesh Maharaj ◽  
Shin-Yi Lin ◽  
Darya O. Mishchuk ◽  
Elizabeth Chin ◽  
...  
Keyword(s):  
High Throughput Sequencing ◽  
Water Soluble ◽  
Resonance Spectroscopy ◽  
Its Gene ◽  
Candidatus Liberibacter ◽  
Liberibacter Asiaticus ◽  
Citrus Leaves ◽  
Citrus Varieties ◽  
And Control ◽  
The Impact

Huanglongbing (HLB) is a severe, incurable citrus disease caused by the bacterium ‘Candidatus Liberibacter asiaticus’ (CLas). Although citrus leaves serve as the site of initial infection, CLas is known to migrate to and colonize the root system; however, little is known about the impact of CLas infection on root metabolism and resident microbial communities. Scions of ‘Lisbon’ lemon and ‘Washington Navel’ orange grafted onto ‘Carrizo’ rootstock were grafted with either CLas-infected citrus budwood or uninfected budwood. Roots were obtained from trees 46 weeks after grafting and analyzed via 1H nuclear magnetic resonance spectroscopy to identify water-soluble root metabolites and high-throughput sequencing of 16S rRNA and ITS gene amplicons to determine the relative abundance of bacterial and fungal taxa in the root rhizosphere and endosphere. In both citrus varieties, 27 metabolites were identified, of which several were significantly different between CLas(+) and control plants. CLas infection also appeared to alter the microbial community structure near and inside the roots of citrus plants. Nonmetric multidimensional scaling (NMDS) and a principal coordinate analysis (PCoA) revealed distinct metabolite and microbial profiles, demonstrating that CLas impacts the root metabolome and microbiome in a manner that is variety-specific.

scholarly journals Effect of Adjuvants on Oxytetracycline Uptake upon Foliar Application in Citrus

Antibiotics ◽  
2020 ◽  
Vol 9 (10) ◽  
pp. 677
Author(s):  
Nabil Killiny ◽  
Faraj Hijaz ◽  
Pedro Gonzalez-Blanco ◽  
Shelley E. Jones ◽  
Myrtho O. Pierre ◽  
...  
Keyword(s):  
Foliar Application ◽  
Foliar Spray ◽  
Bacterial Pathogen ◽  
Water Addition ◽  
Candidatus Liberibacter ◽  
Citrus Huanglongbing ◽  
Liberibacter Asiaticus ◽  
Trunk Injection ◽  
Citrus Leaves ◽  
Target Effects

Recently in Florida, foliar treatments using products with the antibiotics oxytetracycline and streptomycin have been approved for the treatment of citrus Huanglongbing (HLB), which is caused by the putative bacterial pathogen ‘Candidatus Liberibacter asiaticus’. Herein, we assessed the levels of oxytetracycline and ‘Ca. L. asiaticus’ titers in citrus trees upon foliar applications with and without a variety of commercial penetrant adjuvants and upon trunk injection. The level of oxytetracycline in citrus leaves was measured using an oxytetracycline ELISA kit and ‘Ca. L. asiaticus’ titer was measured using quantitative PCR. Low levels of oxytetracycline were taken up by citrus leaves after foliar sprays of oxytetracycline in water. Addition of various adjuvants to the oxytetracycline solution showed minimal effects on its uptake by citrus leaves. The level of oxytetracycline in leaves from trunk-injected trees was higher than those treated with all foliar applications. The titer of ‘Ca. L. asiaticus’ in the midrib of leaves from trees receiving oxytetracycline by foliar application was not affected after four days and thirty days of application, whereas the titer was significantly reduced in oxytetracycline-injected trees thirty days after treatment. Investigation of citrus leaves using microscopy showed that they are covered by a thick lipidized cuticle. Perforation of citrus leaf cuticle with a laser significantly increased the uptake of oxytetracycline, decreasing the titer of ‘Ca. L. asiaticus’ in citrus leaves upon foliar application. Taken together, our findings indicate that trunk injection is more efficient than foliar spray even after the use of adjuvants. Our conclusion could help in setting useful recommendations for the application of oxytetracycline in citrus to improve tree health, minimize the amount of applied antibiotic, reduce environmental exposure, and limit off-target effects.

Temporal Dynamics of Candidatus Liberibacter asiaticus Titer in Mature Leaves from Citrus sinensis cv Valencia Are Associated with Vegetative Growth

2019 ◽  
Vol 113 (2) ◽  
pp. 589-595 ◽  
Author(s):  
Freddy Ibanez ◽  
Lukasz L Stelinski
Keyword(s):  
Vegetative Growth ◽  
Citrus Sinensis ◽  
Temporal Dynamics ◽  
Critical Role ◽  
Insect Vector ◽  
In Planta ◽  
Mature Leaves ◽  
Candidatus Liberibacter ◽  
Liberibacter Asiaticus ◽  
The Impact

Abstract Huanglongbing, a highly destructive disease of citrus species, is associated with a fastidious, gram-negative, phloem-limited bacteria (Candidatus Liberibacter spp.). In Florida, the causative agent of Huanglongbing (HLB) is C. Liberibacter asiaticus (CLas) and it is transmitted by the insect vector, Asian citrus psyllid (Diaphorina citri Kuwayama). Previous investigations have revealed systemic infection of CLas with an erratic and uneven distribution of pathogen in tree phloem. However, previous investigations did not consider the potential impact of plant vegetative growth on presence/absence of CLas in planta. Our objectives were to determine: 1) the effect of vegetative growth of Citrus sinensis (L.) Osbeck cv Valencia on detection of CLas in mature leaves, and 2) the impact of CLas inoculation frequency on progression of CLas titer in citrus leaves through the first year of infection. Temporal dynamics of CLas detection were associated with vegetative flush growth. Surprisingly, there was no difference in CLas titer detected between plants exposed to infected vectors for a one-time 7 d inoculation access period, as compared with plants exposed to continuously breeding CLas-infected insects over the course of an entire year of plant infection. Our results suggest that the CLas bacterium is transported through phloem during annual movement of carbon compounds needed for vegetative plant growth, including transportation from roots to mature leaves. These results highlight the importance of vegetative growth on temporal dynamics of CLas in citrus, and suggest a critical role of the sink-source interaction on presence/absence of CLas in leaves.

scholarly journals Volatile compounds of Persian and Mexican lime associated with HLB (Huanglongbing) symptoms

2018 ◽  
Vol 44 (1) ◽  
pp. 19 ◽  
Author(s):  
Estrella Mendoza-Peña ◽  
Juan Cibrián-Tovar ◽  
Julio Velázquez-González ◽  
Felipe Tafoya-Rangel ◽  
Ausencio Azuara-Domínguez
Keyword(s):  
Early Detection ◽  
Volatile Compounds ◽  
Solid Phase ◽  
Diagnostic Strategy ◽  
Candidatus Liberibacter Asiaticus ◽  
Mass Selective Detector ◽  
Selective Detector ◽  
Candidatus Liberibacter ◽  
Liberibacter Asiaticus ◽  
And Control

The bacterium Candidatus Liberibacter asiaticus is the pathogen that causes the disease known as Huanglongbing (HLB) in citrus. During the course of the disease, the bacterium affects citrus plant phloem tissues, but their leaves remain asymptomatic for HLB for months to years after initial infection. This limits the early detection and control of the bacterium in infected trees. Therefore, in order to design a diagnostic strategy for HLB, the aim of this study was to quantify the abundance and concentration of the volatile compounds released from young shoots of the Persian lemon (Citrus latifolia Tanaka) and the Mexican lemon [Citrus aurantifolia (Christm.) Swingle] with and without HLB symptoms. The volatiles emitted by young shoots were captured by Solid Phase Microextraction (SPME) and analyzed in a gas chromatograph coupled to a mass selective detector (CG / MS). The results clearly indicate that young shoots with and without HLB symptoms released different abundances and concentrations of volatile compounds. The compounds: D-limonene, β-ocimene, and caryophyllene were collected at higher concentrations in the young shoots of both lemon species with HLB symptoms. This result shows the feasibility of designing a strategy for early detection of the disease in different species of lemon through recognition of patterns and concentrations of volatile compounds released from infected trees.

scholarly journals Confirmation of the Sequence of ‘Candidatus Liberibacter asiaticus’ and Assessment of Microbial Diversity in Huanglongbing-Infected Citrus Phloem Using a Metagenomic Approach

2009 ◽  
Vol 22 (12) ◽  
pp. 1624-1634 ◽  
Author(s):  
Heather L. Tyler ◽  
Luiz F. W. Roesch ◽  
Siddarame Gowda ◽  
William O. Dawson ◽  
Eric W. Triplett
Keyword(s):  
Dna Sequences ◽  
High Throughput Sequencing ◽  
Metagenomic Data ◽  
Candidatus Liberibacter Asiaticus ◽  
Metagenomic Dna ◽  
Rna Sequences ◽  
Culture Independent ◽  
Candidatus Liberibacter ◽  
Liberibacter Asiaticus ◽  
Sequencing Platforms

The citrus disease Huanglongbing (HLB) is highly destructive in many citrus-growing regions of the world. The putative causal agent of this disease, ‘Candidatus Liberibacter asiaticus’, is difficult to culture, and Koch's postulates have not yet been fulfilled. As a result, efforts have focused on obtaining the genome sequence of ‘Ca. L. asiaticus’ in order to give insight on the physiology of this organism. In this work, three next-generation high-throughput sequencing platforms, 454, Solexa, and SOLiD, were used to obtain metagenomic DNA sequences from phloem tissue of Florida citrus trees infected with HLB. A culture-independent, polymerase chain reaction (PCR)-independent analysis of 16S ribosomal RNA sequences showed that the only bacterium present within the phloem metagenome was ‘Ca L. asiaticus’. No viral or viroid sequences were identified within the metagenome. By reference assembly, the phloem metagenome contained sequences that provided 26-fold coverage of the ‘Ca. L. asiaticus’ contigs in GenBank. By the same approach, phloem metagenomic data yielded less than 0.2-fold coverage of five other alphaproteobacterial genomes. Thus, phloem metagenomic DNA provided a PCR-independent means of verifying the presence of ‘Ca L. asiaticus’ in infected tissue and strongly suggests that no other disease agent was present in phloem. Analysis of these metagenomic data suggest that this approach has a detection limit of one ‘Ca. Liberibacter’ cell for every 52 phloem cells. The phloem sample sequenced here is estimated to have contained 1.7 ‘Ca. Liberibacter’ cells per phloem cell.

scholarly journals Genome Sequence Resource of ‘Candidatus Liberibacter asiaticus’ strain 9PA From Brazil

Plant Disease ◽  
2021 ◽  
Vol 105 (1) ◽  
pp. 199-201
Author(s):  
P. A. Silva ◽  
J. Huang ◽  
N. A. Wulff ◽  
Z. Zheng ◽  
R. Krugner ◽  
...  
Keyword(s):  
Genome Sequence ◽  
Sweet Orange ◽  
Whole Genome Sequence ◽  
Whole Genome ◽  
Candidatus Liberibacter Asiaticus ◽  
Citrus Production ◽  
Candidatus Liberibacter ◽  
Citrus Huanglongbing ◽  
Liberibacter Asiaticus ◽  
And Control

‘Candidatus Liberibacter asiaticus’, an unculturable α-proteobacterium, is associated with citrus huanglongbing (HLB), a devastating disease threatening citrus production in Brazil and worldwide. In this study, a draft whole-genome sequence of ‘Ca. L. asiaticus’ strain 9PA from a sweet orange (cultivar Pera) tree collected in São Paulo State, Brazil, is reported. The 9PA genome is 1,231,881 bp, including two prophages, with G+C content of 36.7%. This is the first report of a whole-genome sequence of ‘Ca. L. asiaticus’ from Brazil or South America. The 9PA genome sequence will enrich ‘Ca. L. asiaticus’ genome resources and facilitate HLB research and control in Brazil and the world.

scholarly journals Improvement in the Excised Citrus Leaf Assay to Investigate Inoculation of ‘Candidatus Liberibacter asiaticus’ by the Asian Citrus Psyllid Diaphorina citri

Plant Disease ◽  
2017 ◽  
Vol 101 (3) ◽  
pp. 409-413 ◽  
Author(s):  
Laudecir L. Raiol-Junior ◽  
Ana D. B. Baia ◽  
Fernanda Q. B. F. Luiz ◽  
Camila G. Fassini ◽  
Viviani V. Marques ◽  
...  
Keyword(s):  
Quantitative Polymerase Chain Reaction ◽  
Leaf Age ◽  
Infection Process ◽  
Diaphorina Citri ◽  
Candidatus Liberibacter Asiaticus ◽  
Leaf Petiole ◽  
Candidatus Liberibacter ◽  
Liberibacter Asiaticus ◽  
Citrus Leaves ◽  
Time Required

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.

Detection of Liberibacter asiaticus causing Citrus Vein Phloem Degeneration from Siam Citrus leaves (Citrus nobilis var. microcarpa) in Singkawang City plantation, Pontianak, West Kalimantan

2020 ◽  
Vol 14 (3) ◽  
pp. 2
Author(s):  
RAHMAWATI RAHMAWATI ◽  
ILIANA ILIANA ◽  
AGUS RACHMAT ◽  
LATIFFAH ZAKARIA ◽  
MUKARLINA MUKARLINA
Keyword(s):  
16S Rrna ◽  
Protein Gene ◽  
Ribosomal Protein Gene ◽  
Rrna Gene ◽  
Candidatus Liberibacter Asiaticus ◽  
Specific Primers ◽  
West Kalimantan ◽  
Candidatus Liberibacter ◽  
Liberibacter Asiaticus ◽  
Citrus Leaves

The objective of the present study was to detect presence of pathogenic fastidious bacterium, Candidatus Liberibacter asiaticus using PCR from leaves of Siam citrus showing Citrus Vein Phloem Degeneration (CVPD) symptoms in Singkawang City plantation, Pontianak, West Kalimantan, Indonesia. Citrus leaf samples were collected based on visual observation of symptoms showing CVPD infection. Typical symptoms of CVPD include leaf yellowing (chlorosis), vein banding, leaves become stiff, thicker and smaller in size. The pathogenic bacterium, Candidatus Liberibacter asiaticus was detected using two specific primers, namely, OI1/ OI2c amplified 16S rRNA gene and A2/J5, amplified ribosomal  protein gene of the rplKAJL-rpoBC operon (β-operon). PCR amplification detected the presence of 1100 bp band using OI1/ OI2c primers, and 703 bp band using A2/J5 primers from symptomatic Siam citrus leaves. PCR products were not detected from healthy plants serve as control. By using two sets of specific primers to amplify 16S rRNA gene and ribosomal protein gene, Candidatus Liberibacter asiaticus was detected in symptomatic Siam citrus leaves in Singkawang City, Pontianak, Indonesia. Detection of the bacterial pathogen causing CVPD is important to prevent spreading of the disease which could affect production of citrus fruits.

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