A protocol for successful transmission of ‘Candidatus Liberibacter asiaticus’ from citrus to citrus using Diaphorina citri

2021 ◽  
Author(s):  
Silvio Aparecido Lopes ◽  
Juan Camilo Cifuentes-Arenas
Keyword(s):  
Ambient Air ◽  
Diaphorina Citri ◽  
Candidatus Liberibacter Asiaticus ◽  
Mature Leaves ◽  
Air Temperatures ◽  
Tree Canopies ◽  
Candidatus Liberibacter ◽  
Liberibacter Asiaticus ◽  
Time Required ◽  
Transmission Studies

A protocol to successfully transmit the huanglongbing (HLB) pathogen, ‘Candidatus Liberibacter asiaticus’ (Las), from citrus-to-citrus plants using the Asian citrus psyllid (ACP), and an alternative way to help growers control ACP are proposed. Best results were obtained when pathogen acquisition by adults reared on fully symptomatic Las-positive plants, latency, and inoculation, occurred at ambient air temperatures ranging from 24 to 28°C, and by confining single infective adult ACP for seven days on soft newly developing vegetative shoots (stages v2 to v4). No infection resulted from confinement of infective ACP adults on mature leaves (stage v6). Under the described conditions, single ACP adults could successfully transmit Las to an average of 56.5% (35 to 83%) of plantlets with v2 to v4 shoots growing in 0.3 L tubes, and to 80.5% (76 to 86%) of plants with v2 to v4 shoots growing in 4.7 L pots. The use of single insects and plantlets reduces labor, space and other resources required to undertake transmission tests. It also reduces time required for transmission studies and should help accelerate research on HLB. The results were used to develop an index for favorability to infection (IFI) to determine orchard vulnerabilities to Las. The IFI is based on the heterogeneous population of new shoots that occurs on tree canopies and may offer alternative or complementary alternatives to the laborious and costly insect surveys currently used in most instances to determine threshold levels for insecticide applications.

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.

scholarly journals ‘Candidatus Liberibacter asiaticus’ Titers in Citrus and Acquisition Rates by Diaphorina citri Are Decreased by Higher Temperature

Plant Disease ◽  
2013 ◽  
Vol 97 (12) ◽  
pp. 1563-1570 ◽  
Author(s):  
S. A. Lopes ◽  
F. Q. B. F. Luiz ◽  
E. C. Martins ◽  
C. G. Fassini ◽  
M. C. Sousa ◽  
...  
Keyword(s):  
Quantitative Polymerase Chain Reaction ◽  
Disease Spread ◽  
Diaphorina Citri ◽  
Candidatus Liberibacter Asiaticus ◽  
Ambient Temperatures ◽  
Air Temperatures ◽  
Temperature Regimes ◽  
Candidatus Liberibacter ◽  
Liberibacter Asiaticus ◽  
Higher Temperature

‘Candidatus Liberibacter asiaticus’ is the most prevalent Liberibacter sp. associated with huanglongbing (HLB) in Brazil. Within São Paulo state (SP), HLB has spread more rapidly to and reached higher incidence in regions with relatively mild (cooler) summer temperatures. This suggests that climate can influence disease spread and severity. ‘Ca. L. asiaticus’ titers on soft, immature leaves from infected ‘Valencia’ sweet orange plants exposed to different temperature regimes and adult Diaphorina citri fed for 48 h on these plants for ‘Ca. L. asiaticus’ acquisition were determined by quantitative polymerase chain reaction in two experiments. The first experiment included plants with three levels of infection, three incubation periods (IPs), and air temperatures favorable (14.6 to 28°C) and unfavorable (24 to 38°C) to ‘Ca. L. asiaticus’. The second included plants with severe late-stage infections, 10 IPs (based on 3-day intervals over 27 days), and three air temperature regimes (12 to 24, 18 to 30, and 24 to 38°C). Overall, ‘Ca. L. asiaticus’ titers and the percentages of ‘Ca. L. asiaticus’-positive psyllids were lower in plants maintained at the warmer temperature regime (24 to 38°C) than in plants maintained in the cooler regimes. The results suggest that the lower incidence and slower spread of ‘Ca. L. asiaticus’ to warmer regions of SP are related to the influence of ambient temperatures on titers of ‘Ca. L. asiaticus’ in leaves.

Probing the application of OmpA-derived peptides to disrupt the acquisition of ‘Candidatus Liberibacter asiaticus’ by Diaphorina citri

2021 ◽  
Author(s):  
Marcus Vinicius Merfa e Silva ◽  
Eduarda Regina Fischer ◽  
Mariana de Souza e Silva ◽  
Carolina Sardinha Francisco ◽  
Helvécio Coletta-Filho ◽  
...  
Keyword(s):  
Control Strategies ◽  
Insect Vector ◽  
Diaphorina Citri ◽  
Candidatus Liberibacter Asiaticus ◽  
In Planta ◽  
Host Interaction ◽  
Vector Population ◽  
Candidatus Liberibacter ◽  
Liberibacter Asiaticus ◽  
Bacterial Outer Membrane

Huanglongbing (HLB) is currently the most devastating disease of citrus worldwide. Both bacteria ‘Candidatus Liberibacter asiaticus’ (CLas) and ‘Ca. Liberibacter americanus’ (CLam) are associated with HLB in Brazil, but with a strong prevalence of CLas over CLam. Conventionally, HLB management focuses on controlling the insect vector population (Diaphorina citri; also known as Asian citrus psyllid – ACP) by spraying insecticides, an approach demonstrated to be mostly ineffective. Thus, development of novel more efficient HLB control strategies is required. The multifunctional bacterial outer membrane protein OmpA is involved in several molecular processes between bacteria and their hosts and has been suggested as a target for bacterial control. Curiously, OmpA is absent in CLam in comparison to CLas, suggesting a possible role on host-interaction. Therefore, in the current study, we have treated ACPs with different OmpA-derived peptides aiming to evaluate the acquisition of CLas by the insect vector. Treatment of psyllids with 5 µM of Pep1, Pep3, Pep5 and Pep6 in artificial diet significantly reduced the acquisition of CLas, while increasing the concentration of Pep5 and Pep6 to 50 µM abolished this process. In addition, in planta treatment with 50 µM of Pep6 also significantly decreased the acquisition of CLas and sweet orange plants stably absorbed and maintained this peptide for as long as three months post the final application. Together, our results demonstrate the promising use of OmpA-derived peptides as a novel biotechnological tool to control CLas.

Population diversity of “Candidatus Liberibacter asiaticus” and Diaphorina citri in Sichuan: A case study for Huanglongbing monitoring and interception

Plant Disease ◽  
2021 ◽  
Author(s):  
Xuejin Cui ◽  
Kehong Liu ◽  
Jie Huang ◽  
Shimin Fu ◽  
Qingdong Chen ◽  
...  
Keyword(s):  
Rural Areas ◽  
Urban Areas ◽  
Control Strategies ◽  
Population Diversity ◽  
Mitochondrial Genomes ◽  
Diaphorina Citri ◽  
Candidatus Liberibacter Asiaticus ◽  
Multiple Sources ◽  
Candidatus Liberibacter ◽  
Liberibacter Asiaticus

Citrus Huanglongbing (HLB) is present in 10 provinces in China and is associated with “Candidatus Liberibacter asiaticus” (CLas), which is transmitted by the Asian citrus psyllid (Diaphorina citri, ACP). To date, HLB and ACP have expanded to Yibin city of Sichuan Province, posing an imminent threat to the citrus belt of upper and middle reach of Yangtze River, an important late maturing citrus-producing area in China. To understand the epidemiological route of CLas and ACP in newly invaded regions of Sichuan and thereby better establish an HLB-interception zone ranging from Leibo to Yibin, we evaluated the molecular variability of 19 CLas draft genomes from citrus or dodder (Cuscuta campestris). They include three type-specific prophage loci, three variable numbers of tandem repeat (VNTR) loci, a miniature inverted-repeat transposable element (MITE) types, and population diversity of 44 ACP mitochondrial genomes. The results indicated that CLas isolates in the newly invaded area (Pingshan) were more diverse than those in the HLB endemic areas (Leibo and Ningnan). Phylogenetic analysis based on mitochondrial genomes demonstrated that ACPs in Leibo, Pingshan and Xuzhou (rural areas) represent a new group (MG4), distinguished by the three unique SNPs in cox1, nad4 and cytb. However, the ACPs sampled from the urban areas of Cuiping and Xuzhou belonged to the southeastern China group (MG2-1). Altogether, our study revealed multiple sources of ACP and CLas in the HLB-interception zone and proposed their transmission route. This study contributes to the formulation of precise HLB prevention and control strategies in the HLB-interception zone in Sichuan and could be useful for HLB management efforts in other regions.

A Review of Huanglongbing (Citrus Greening) Management in Citrus Using Nutritional Approaches in China

2011 ◽  
Vol 12 (1) ◽  
pp. 24 ◽  
Author(s):  
Yulu Xia ◽  
Gecheng Ouyang ◽  
Ronald A. Sequeira ◽  
Yu Takeuchi ◽  
Ignacio Baez ◽  
...  
Keyword(s):  
Nutrient Management ◽  
Field Survey ◽  
Cultural Practices ◽  
Sweet Orange ◽  
Asian Citrus Psyllid ◽  
Diaphorina Citri ◽  
Candidatus Liberibacter Asiaticus ◽  
Candidatus Liberibacter ◽  
Citrus Maxima ◽  
Liberibacter Asiaticus

The Asian form of huanglongbing (HLB) is caused by ‘Candidatus Liberibacter asiaticus (Las),’ a phloem-limited bacterium transmitted by the Asian citrus psyllid, Diaphorina citri Kuwayama. Nutrient management, together with other cultural practices such as pruning and irrigation, for mitigation of the disease has been practiced in China for many years. Our literature review, field survey, and interviews with Chinese scientists and growers indicate that these cultural practices were generally ineffective for the disease management. However, a nutritional approach in conjunction with other cultural practices such as irrigation can maintain grove productivity for a certain time depending on the type of citrus species/cultivars, the age of the trees, the propagation method of the plants, the Asian citrus psyllid (ACP) (Diaphorina citri Kuwayama) population, and other factors. Symptomatic mature pommelo (Citrus maxima Merr) and sweet orange (C. sinensis L. Osbeck) plants can commonly survive and maintain a certain level of productivity for an additional 4 to 5 years, even longer assuming vigorous ACP control. Accepted for publication 27 June 2011. Published 3 October 2011.

scholarly journals Tracing Penicillin Movement in Citrus Plants Using Fluorescence-Labeled Penicillin

Antibiotics ◽  
2019 ◽  
Vol 8 (4) ◽  
pp. 262
Author(s):  
Nabil Killiny ◽  
Pedro Gonzalez-Blanco ◽  
Yulica Santos-Ortega ◽  
Fuad Al-Rimawi ◽  
Amit Levy ◽  
...  
Keyword(s):  
Vascular Tissue ◽  
Vascular System ◽  
Citrus Greening ◽  
Application Site ◽  
Asian Citrus Psyllid ◽  
Diaphorina Citri ◽  
Candidatus Liberibacter Asiaticus ◽  
Citrus Industry ◽  
Candidatus Liberibacter ◽  
Liberibacter Asiaticus

Huánglóngbìng (HLB), citrus greening, is one of the most destructive diseases of citrus plants worldwide. In North America, HLB is caused by the phloem-limited bacterium Candidatus Liberibacter asiaticus and is transmitted by the Asian citrus psyllid, Diaphorina citri. No cure exists at present, and the use of antibiotics for the control of HLB has gained interest due to the significant losses to the citrus industry. Because of unsatisfactory results when using foliar applications of antibiotics, concerns were raised regarding the uptake and translocation of these materials within trees. We, therefore, investigated a method that allows us to study the movement of antibiotic materials in citrus plants. Herein, we utilized a fluorescence-labeled penicillin, BOCILLIN™ FL-Penicillin (FL-penicillin), to study the uptake and translocation of penicillin in citrus plants. FL-penicillin was applied by puncture to the stem of young citrus seedlings and was traced by using fluorescence microscopy. After application, we detected FL-penicillin in the leaves and in the stem xylem and phloem tissues above and below the application site in both intact and partially bark-girdled citrus seedlings, indicating that it is easily taken up and transported through the plant vascular system. In addition, we detected FL-penicillin in the gut of D. citri, which were allowed to feed on the treated plants, suggesting translocation of this molecule into the vascular tissue. We propose that the use of fluorescent-labeled molecules could be an effective tool for understanding the uptake and translocation of antibiotics and other macromolecules in plants and insects.

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