Narrow host range phages associated with citrus canker lesions in Florida and Argentina

Bacteriophages were isolated from naturally infected citrus canker lesions from diverse locations in Florida and Argentina and characterized for host range using a world-wide collection of Xanthomonas citri subsp. citri (Xcc) strains. Sixty-seven bacteriophages isolated from citrus canker lesions in Florida (37 bacteriophages) and Argentina (30 bacteriophages) revealed little diversity. All 30 phages isolated from four locations in Argentina had identical host ranges (group ARG), while 37 phages from Florida made up two groups (FLA and FLB). ARG and the 31 FLA phages produced clear plaques and had nearly identical host ranges as phage CP2 of Japan in that they only reacted with typical A strains and none of the atypical A strains (A^* and A^W) or other Xanthomonas spp. FLB phages had a different host range from the other strains and produced turbid plaques. We used phage typing, fatty acid analysis and riboprinter analysis to classify citrus-associated xanthomonads. Phage typing using 12 phages isolated from Xcc, X. fuscans subsp. aurantifolii (Xfa), X. alfalfae subsp. citrumelonis (Xacm), and other sources proved useful for classifying all major Xcc pathotypes and/or strains (A, A*, Miami (MI), Manatee (MA) and Wellington (A^W)), as well as B and C types of Xfa. X. citri subsp. citri strains from a worldwide collection were diverse in phage susceptibility. The majority of Xcc strains, which originated from different regions of the world and which were typical “A” pathotype strains based on pathogenicity characteristics, was sensitive to most phages (including CP2, FLA and ARG), and had nearly identical phage sensitivity profiles. MA strains were quite unique in that they reacted with none of the phages; furthermore, they were different from the putative progenitor MA strain, ATCC 49118, which reacted with a group of phages. Fatty acid analysis revealed considerable variation in Xcc-A, Xfa-B, Xfa-C and Xacm strains. Using riboprinter analysis, we identified a unique riboprinter pattern for strains isolated from an etrog tree (Citrus medica) in Florida that were “A” pathotype strains based on pathogenicity characteristics. Phage typing and fatty acid analysis were useful in corroborating that the etrog strains represent a unique new Xcc strain in Florida.     

AR模型在柑桔溃疡病测报中的应用

 柑桔溃疡病发生程度的预测是进行该病治理的重要依据。本研究用自回归方法对2004年3月12日~2005年4月13日期间调查的病情原始数据进行标准化、平稳化处理以及模型的参数估计和拟合检验,所得AR (14)模型可以精确预测溃疡病发生趋势。     

柑橘溃疡病检疫与防治

细菌性溃疡病是严重危害世界柑橘产业的重大检疫性病害之一,柑橘溃疡病引起落叶、枯枝和落果,溃疡病斑导致果品质量降低,影响外贸出口。世界各国长期以来对病害采取严格苗木检疫、疫区病树铲除、零星病害药剂防控的综合治理措施;新近美国农业部推出"柑橘健康种植行动计划";2007年7月中国农业部正式启动"柑橘非疫区建设和维护"项目,总体目标在于防控柑橘溃疡病的发生和传播,确保柑橘产业的安全。     

The Horsfall-Barratt scale and severity estimates of citrus canker

Citrus canker assessment data were used to investigate effects of using the Horsfall-Barratt (H-B) scale to estimate disease compared to direct estimation to the nearest percent. Twenty-eight raters assessed each of two-hundred infected leaves (0–38% true diseased area). The data were converted to the H-B scale. Correlation (r) showed that direct estimates had higher inter-rater reliability compared to H-B scaled data (r = 0.75 and 0.71 for direct estimates and H-B scaled data, respectively). Lin’s concordance correlation (LCC, ρ _c ) analysis showed individual rater estimates by direct estimation had better agreement with true values compared to H-B scaled data. The direct estimates were more precise compared to H-B scaled data (r = 0.80–0.95 and 0.61–0.90, respectively), but measures of generalised bias or accuracy (C _b ) were similar for both methods (0.38–1.00). Cumulative mean disease and cumulative variance of the means were calculated for each rater on a leaf-by-leaf basis. Direct estimates were closer to the true severity 59.5% of the time, and to the cumulative true sample mean 53.7% of the time, and to the cumulative true sample mean variance 63.6% of the time. Estimates of mean severity for each leaf based on estimates by 3, 5, 10, 20 and 28 raters were compared to true disease severity. LCC showed that rater-means based on more raters had better agreement with true values compared to individual estimates, but H-B scale data were less precise, although with means based on ≥ 10 raters, agreement was the same for both assessment methods. Magnitude and dispersion of the variance of the means based on H-B scaled data was greater than that by direct estimates. H-B scaling did not improve reliability, accuracy or precision of the estimate of citrus canker severity compared to direct visual estimation.     

Ability of governments to take actions to confront incursions of diseases – a case study: citrus canker in Florida

When governments take actions under their police powers to prevent incursions of diseases, they may damage or destroy private property and a question arises of who should pay for property losses. A court in Florida concluded that the state needed to pay for property destroyed under a citrus canker eradication programme. Because this interpretation of the Florida Constitution’s Just Compensation Clause makes it more difficult to administer a successful eradication programme, governments and scientists, working with industry, may want to develop a more comprehensive accounting of scientific data and procedures that justify an eradication programme’s actions. This paper identifies three problems regarding Florida’s efforts to eradicate citrus canker so that scientists, affected industries, and governments can employ an objective reassessment mechanism to support their actions without legal interference.     

不同柑桔品种对柑桔溃疡病抗病能力的测定

柑桔溃疡病为国内外检疫对象,危害柑桔苗木、幼树和成年树的叶片、果实、树梢等,引起落叶、落果、枯梢、削弱树势,是柑橘的重要病害.不同的柑桔品种对该病的抗病性差异程度较大,为了明确不同种类的柑桔品种对该病的抗病性,我们进行了柑桔不同品种对其抗性能力的测定.     

柑桔溃疡病发生特点及综合防治对策

柑桔溃疡病(Xanthomnas campestris pv.citri(Hasse)Dye是国际、国内植物检疫对象。目前该病在我国14个省(市)有发生,发生面积约8.7万hm2,虽然发生面积小,仅占柑桔种植面积的4.38%,但是对溃疡病柑桔生产和贸易造成了重大影响。柑桔是江西省农业的支柱产业,控制柑桔溃疡病的发生危害,确保柑桔生产安全,提高产量和果品质量,使其具有国内外市场竞争力,是当前江西省植物保护工作的重要内容之一。     

Copper rate and spray interval for joint management of citrus canker and citrus black spot in orange orchards

European Journal of Plant Pathology - Citrus canker and citrus black spot are two of the most important diseases in the São Paulo (SP) citrus belt, Brazil. Canker control was based on an...     

柑桔溃疡病内生拮抗细菌Bc51的研究

 从中国南宁柑桔叶片中分离到对柑桔溃疡病菌有拮抗作用的细菌菌株Bc51。根据16S rDNA序列同源性、形态学特征和生理生化反应,将该菌株鉴定为洋葱伯克霍尔德氏菌(Burkholderia cepacia)。温度、pH值和培养基对洋葱伯克霍尔德氏菌抑制柑桔溃疡病菌生长的能力有显著影响。在温室测试中,观察到60.3%柑桔溃疡病斑的形成受到洋葱伯克霍尔德氏菌抑制。连续8次在人工培养基上转代培养,洋葱伯克霍尔德氏菌对柑桔溃疡病菌生长的抑制力没有发生明显改变。洋葱伯克霍尔德氏菌对植物病原菌有较宽的抑菌谱,表明除柑桔溃疡病外,该菌对其它作物病害的防治也具有潜在的应用价值。     

Detached leaf inoculation of germplasm for rapid screening of resistance to citrus canker and citrus bacterial spot

A detached leaf protocol for rapid screening of germplasm for resistance to citrus canker (Xanthomonas citri subsp. citri, Xcc) and citrus bacterial spot (Xanthomonas alfalfae subsp. citrumelonis, Xac) was developed to evaluate limited quantities of leaf material. Bacterial inocula of Xcc or Xac at 10⁴, 10⁵, or 10⁸ cfu ml⁻¹ were injection-infiltrated into the abaxial surface of disinfested, immature leaves of susceptible and resistant genotypes. Inoculated detached leaves were placed on the surface of 0.5% water agar plates and incubated at 28°C under a 12 h photoperiod. Likewise, inocula were infiltrated into attached leaves of greenhouse plants. At high inoculum concentrations of Xcc or Xac (10⁸ cfu ml⁻¹), resistant cultivars of kumquat developed a hypersensitive-like reaction within 3 days post inoculation (dpi). At 10⁵ cfu ml⁻¹, populations 14 dpi were <10⁴ per inoculation site. In canker-susceptible Citrus spp. (‘Duncan’ grapefruit and ‘Rough’ lemon), water-soaked areas occurred by 3 dpi and typical canker lesions developed by 7 to14 dpi. Concentration of Xcc recovered from inoculation sites was approximately 10⁵ cfu ml⁻¹ by 14 dpi. In citrus bacterial spot-susceptible citrus (‘Swingle’ citrumelo and grapefruit), symptoms developed within 7 dpi. Populations of Xac after inoculation at 10⁵ cfu ml⁻¹ were comparable to Xcc in susceptible hosts 14 dpi (>10⁵). The detached leaf assay is useful for the characterization and differentiation of lesion phenotype for each Xanthomonas pathogen permitting rapid screening of germplasm resistance based on the quantification of number of lesions and bacterial concentration.     

Over-expression of the citrus gene CtNH1 confers resistance to bacterial canker disease

Citrus canker is a devastating disease, caused by Xanthomonas axonopodis pv. citri (Xac). It is well established that the NPR1 gene plays a pivotal role in systemic acquired resistance (SAR) in Arabidopsis. In this study, we report the isolation and characterization of an NPR1 homolog from citrus, namely Citrus NPR1 homolog 1 (CtNH1). Sequence alignment and phylogenetic analysis indicate that CtNH1 is closely-related to the Arabidopsis NPR1 gene and its orthologs from rice, grapevine, and cacao. When over-expressed in citrus, CtNH1 confers resistance to Xac and leads to constitutive expression of the pathogenesis-related (PR) gene chitinase 1 (Chi1), suggesting that CtNH1 is orthologous to NPR1.     

Enhanced resistance to citrus canker in transgenic sweet orange expressing the sarcotoxin IA gene

Citrus canker, caused by the bacterial pathogen Xanthomonas citri subp. Citri (Xcc), is a serious disease reported in most citrus-producing areas around the world. Although different levels of field resistance to citrus canker have been reported in sweet oranges, they are usually not sufficient to provide adequate control of the disease. Ectopic over-expression of antibacterial genes is one of the potential strategies to increase plant resistance to bacterial diseases. Previous in vitro results showed that sarcotoxin IA, an antimicrobial peptide isolated from the flesh fly (Sarcophaga peregrina), can be efficient to control different plant pathogenic bacteria, including Xcc. Transgenic “Pera” sweet orange (Citrus sinensis [L.] Osbeck) plants constitutively expressing the sarcotoxin IA peptide fused to the PR1a signal peptide from Nicotiana tabacum for secretion in the intercellular space were obtained by Agrobacterium-mediated transformation using thin sections of mature explants. Citrus canker resistance evaluation in leaves of transgenic and non-transgenic plants was performed through inoculations with Xcc by infiltration and spraying. The Xcc population was up to 2 log unit lower in leaves of the transgenic plants compared to those of non-transgenic controls. Incidence of canker lesions was significantly higher in non-transformed controls (>10 lesions/cm²) than in the transgenic plants (<5 lesions/cm²) after injection infiltration or spraying with Xcc inoculum. Accumulation of sarcotoxin IA peptide in sweet orange tissue did not cause any deleterious effects on the growth and development of the transgenic plants, indicating this approach is suitable to provide resistance to citrus canker.     

柑橘溃疡病菌检测方法及其在检疫中的优选

采用常规检测法、免疫斑点法、酶联免疫检测法和PCR检测法对柑橘溃疡病菌纯培养和柑橘叶片提取液进行平行测定。结果表明,对柑橘叶片提取液的检测,4种检测法检出的样品带菌率分别为11.12%、27.22%、18.69%、20.31%;对柑橘溃疡病菌纯培养的检测,4种方法检测结果一致。本实验同时比较了几种方法的灵敏度、特异性和稳定性,以及不同方法在各级检验检疫中的适应性。     

正确认识柑桔溃疡病对赣南脐橙生产的影响

赣南具有得大独厚的脐橙生产条件，赣州市委、市政府把脐橙生产作为“山上再造一个新赣南”的支柱产业来抓，脐橙种植面积不断扩大，至2003年已经达到5．47万hm^2。但是，随着脐橙面积的增加，柑桔溃疡病也呈蔓延之势，已严重制约赣南脐橙的可持续发展，进行柑桔溃疡病的综合治理迫在眉睫。     

Differentiation of citrus bacterial canker strains in Korea by host range,rep-PCR fingerprinting and 16S rDNA analysis

Soil-borne cereal mosaic virus (SBCMV) causes a severe disease in susceptible cultivars of winter wheat. The virus is vectored by the soil-borne protist Polymyxa graminis. Experiments were conducted to investigate whether SBCMV RNA2 could persist in seed from SBCMV-infected susceptible cultivars of winter wheat. Over 7,000 seedlings were generated from seed collected from two cultivars of SBCMV-infected winter wheat. Seedlings were grown in a glasshouse compartment and batch tested for the presence of SBCMV using real-time RT-PCR. The majority of batches tested positive for SBCMV, indicating an RNA2 transmission rate of 1.8–9.4% in wheat. The presence of the virus was confirmed by amplifying and sequencing a larger (400 bp) fragment of viral RNA2 in a sub-set of the seedlings testing positive by real-time RT-PCR. Root extracts from this sub-set tested negative for P. graminis using real-time PCR. The implications for disease epidemiology of this virus are discussed. The authors are British Civil Servants and as such their work is subject to British Crown Copyright. This means the exclusive copyright for the article cannot be transferred.     

Insecticide resistance in field populations of Asian citrus psyllid in Florida

BACKGROUND: Asian citrus psyllid (ACP), Diaphorina citri, is a major pest of citrus because it vectors the putative causal agent of huanglongbing disease. Insecticides are currently the basis of psyllid management programs, and the number of annual insecticide applications has increased significantly. In this paper, a series of investigations of insecticide resistance among field populations of adult and immature ACP in Florida is described. RESULTS: In 2009, the highest level of resistance for adult ACP, as compared with the laboratory susceptible (LS) population, was found with imidacloprid with an LD₅₀ resistance ratio (RR₅₀) of 35 in one population. This was followed by chlorpyriphos (RR₅₀ = 17.9, 13.3, 11.8 and 6.9), thiamethoxam (RR₅₀ = 15 and 13), malathion (RR₅₀ = 5.4 and 5.0) and fenpropathrin (RR₅₀ = 4.8). In 2010, mortality of adults from all five sites sampled was lower than with the LS population at three diagnostic concentrations of each insecticide tested. Among nymph populations, indications of resistance were observed with carbaryl (RR₅₀ = 2.9), chlorpyriphos (RR₅₀ = 3.2), imidacloprid (RR₅₀ = 2.3 and 3.9) and spinetoram (RR₅₀ = 4.8 and 5.9). General esterase, glutathione S‐transferase and monooxygenase levels were also elevated in field‐collected adult and nymph ACP as compared with the LS population. CONCLUSION: The present results suggest that varying levels of insecticide susceptibility exist in ACP populations across the citrus‐growing areas of Florida. Increased levels of detoxifying enzymes in these populations may partially explain these differences. The present results indicate that insecticide resistance may become an emerging problem for ACP control if effective resistance management is not practiced. Copyright © 2011 Society of Chemical Industry     

Bacteria from the citrus phylloplane can disrupt cell–cell signalling in Xanthomonas citri and reduce citrus canker disease severity

Xanthomonas citri subsp. citri (Xcc) is the causal agent of citrus canker, a disease that affects almost all types of citrus crops. Production of particular Xcc pathogenicity factors is controlled by a gene cluster rpf, which encodes elements of a cell–cell communication system called quorum sensing (QS), mediated by molecules of the diffusible signal factor (DSF) family. Interference with cell–cell signalling, also termed quorum quenching, either by signal degradation or over‐production, has been suggested as a strategy to control bacterial disease. In this study, three bacterial strains were isolated from citrus leaves that displayed the ability to disrupt QS signalling in Xcc. Pathogenicity assays in sweet orange (Citrus sinensis) showed that bacteria of the genera Pseudomonas and Bacillus also have a strong ability to reduce the severity of citrus canker disease. These effects were associated with alteration in bacterial attachment and biofilm formation, factors that are known to contribute to Xcc virulence. These quorum‐quenching bacteria may represent a highly valuable tool in the process of biological control and offer an alternative to the traditional copper treatment currently used to treat citrus canker disease.     

Bacterial antagonists and their cell-free cultures efficiently suppress canker disease in citrus lime

Journal of Plant Diseases and Protection - Proliferation of citrus canker disease caused by Xanthomonas citri subsp. citri (Xcc) has increasingly become a serious threat and has resulted in a...     

Use of phages for identifying the citrus canker bacterium Xanthomonas campestris pv. citri in Taiwan

Phages CP115 and CP122, which were isolated from canker lesions on grapefruit and Liucheng sweet orange, respectively, showed a high degree of specificity with respect to lysis of test bacterial strains. When used jointly, they lysed 135 (97·8%) out of 138 Xanthomonas campestris pv. citri strains isolated from the canker lesions on leaves, twigs, and fruits of various citrus species, cultivars, and hybrids grown throughout Taiwan, but they did not lyse other X. campestris pathovars and other phytopathogenic bacteria, nor other bacteria isolated from soil, clinical or environmental samples. Of 252 CP115/CP122-sensitive and 78 CP115/CP122-resistant bacterial strains with colony characteristics typical of or similar to those of X. campestris pv. citri , isolated from canker lesions of various citrus plants in diverse growing regions in Taiwan, 250 (99·2%) and 76 (97·4%) strains were pathogenic and non-pathogenic, respectively, when inoculated into Liucheng sweet orange or Mexican lime. Thus, phages CP115 and CP122, when used jointly, appear to be applicable for identifying X. campestris pv. citri in Taiwan.