A new haplotype of “Candidatus Liberibacter solanacearum” identified in the Mediterranean region

“Candidatus Liberibacter solanacearum”, a phloem-limited and Gram-negative bacterium that is spread from infected to healthy plants by psyllid insect vectors, is an economically important pathogen of solanaceous and carrot crops in the Americas, New Zealand and Europe. Three haplotypes of “Ca. L. solanacearum” have previously been described, two (LsoA and LsoB) in relation to solanaceous crops in the Americas and New Zealand and the third (LsoC) to carrots in Finland. Herein, we describe a fourth haplotype of this ‘Candidatus Liberibacter’ species (LsoD), also associated with carrots, but from Spain and the Canary Islands and vectored by the psyllid Bactericera trigonica. In addition, LsoC was confirmed in carrot and psyllid samples recently collected from Sweden and Norway. Phylogenetic analysis of the 16S rRNA gene suggests that two of the haplotypes, one in the Americas and the other in northern Europe are closer to each other in spite of a large geographic separation and host differences. Furthermore, during this study, potatoes with symptoms of zebra chip disease recently observed in potato crops in Idaho, Oregon and Washington states were analyzed for haplotype and were found to be positive for LsoA. This liberibacter haplotype was found in psyllids associated with the diseased potato crops as well. This finding contrasts with an earlier report of LsoB from psyllids in Washington which came from a laboratory colony originally collected in Texas.     

Molecular diversity of ‘Candidatus Phytoplasma pyri’ isolates in Slovenia

A European quarantine organism ‘Candidatus Phytoplasma pyri’ causing devastating pear decline disease has been reported to affect pear trees in several European countries. In this study a multilocus sequence analysis was successfully used to gain detailed insight into the molecular diversity of thirty closely related ‘Candidatus Phytoplasma pyri’ isolates from different orchards in Slovenia. Among three genomic regions analyzed, the 16S/23S rRNA intergenic spacer region was the most conserved among Slovenian isolates with 99.7 % sequence identity, yielding only three distinct genotypes. On the other hand, five different genotypes were detected when analyzing secY and aceF genomic regions that shared sequence identity of 94.8 and 97.2 %, respectively. Six of the detected genotypes, specifically four in the secY region and one in each of the two other analyzed genomic regions, were unique for Slovenia. At least eight different haplotypes were found with multilocus sequence analysis, indicating high molecular diversity among Slovenian ‘Ca. P. pyri’ isolates. Haplotypes were clustered into two major clusters, separated by at least 45 mutations. No connection was established between haplotype occurrence and cultivar type.     

Studies on seed transmission of “Candidatus Liberibacter solanacearum” in pepper and its impact on plant emergence

“ Candidatus Liberibacter solanacearum” (Lso), transmitted by the potato psyllid (Bactericera cockerelli), is the causal agent of potato zebra chip, but can also infect other solanaceous plants, including peppers. Studies were conducted to investigate whether Lso could be transmitted to the next generation of plants through seeds from infected pepper plants. In 2014, jalapeno pepper plants were infested with psyllids carrying a mixture of Lso A and B (AB) at the AgriLife Research Station at Bushland. The study was again conducted in 2019 and pepper plants were infested with psyllids carrying Lso B or Lso AB. In each of the studies, noninfested plants served as controls. At harvest, fruits were collected and tested for the presence of Lso using quantitative PCR. Seeds from infected fruits were then tested for Lso. Overall, the percentage of seeds that tested positive for Lso ranged from 33% to 70%. However, Lso detection in embryos ranged only from 0% to 8%. Seed samples from Lso-positive fruits were planted in the greenhouse to determine the impact of Lso on emergence and the incidence of Lso in emerged plants. Although plant emergence differed between some of the seeds obtained from Lso-positive and -negative fruits, the overall impact of Lso on plant emergence was not consistent. However, of the 182 plants that emerged from seeds collected from infected fruits, none was positive for Lso, suggesting that seeds are unlikely to serve as sources for new Lso infections and their impact on disease epidemiology is negligible.     

Prophage region and short tandem repeats of “Candidatus Liberibacter asiaticus” reveal significant population structure in China

Whole-genome sequencing of “Candidatus Liberibacter asiaticus” (Las) indicated some polymorphic gene regions enabling the molecular characterization of this bacterium. Although the population diversity of Las in China has been previously studied, no reports have used a combination of a prophage region and short tandem repeat (STR) loci for phylogenetic relationship characterization. In this study, we investigated the genetic diversity and structure of 667 Las strains from nine provinces in southern China using multiple genetic loci including three type-specific prophage loci, two STR loci, and a miniature inverted-repeat transposable element (MITE) region. The results indicated that the genetic diversity varied according to the gene loci used. The prophage regions, including the MITEs, revealed significant genetic differences of Las populations in Yunnan and Guizhou from other provinces, while the STR loci only indicated a difference of Las population in Guizhou from those of other provinces. In particular, the Las population shown to be diverse in Jiangxi at the CLIBASIA_01215_STR locus was not diverse when measured using the other loci. Considering all loci, the Las populations from Yunnan and Guizhou were different, those from Guangdong and Guangxi were complex, while the Las population in Jiangxi was comparatively simple. Las populations from five neighbouring provinces, including Guangdong, Guangxi, Hunan, Fujian, and Jiangxi were grouped in a big cluster. The Las population structure changed with age according to the prophage type. This study evaluated the method used for studying the molecular diversity of Las populations and provided more detailed information on the geographical origin and epidemic trend of Chinese Las populations.     

Correction to: Haplotypes of ‘Candidatus Liberibacter solanacearum’ identified in Umbeliferous crops in Spain

European Journal of Plant Pathology -     

Molecular analysis of ‘Candidatus Phytoplasma aurantifolia’ associated with phytoplasma diseases in Myanmar

During 2010 and 2011, typical phytoplasma disease symptoms such as little leaves, phyllody and witches’ brooms were observed on black gram, green gram, long bean, shaggy button weed and sesame plants from different regions of Myanmar. The symptomatic samples were analyzed by PCR using universal phytoplasma primers and characterized by sequencing 16S rRNA, ribosomal protein and translocase protein genes. Based on sequence and phylogenetic analysis of the three genes, the phytoplasmas associated with those plants belonged to members of ‘Candidatus Phytoplasma aurantifolia’. To our knowledge, black gram and shaggy button are new hosts for ‘Ca. P. aurantifolia’.     

‘Candidatus Liberibacter solanacearum’ detected in Trioza urticae using suction trap-based monitoring of psyllids in Germany

Journal of Plant Diseases and Protection - Psyllids are small, phloem-feeding insects. Several species are vectors of economically important pathogens, such as ‘Candidatus Liberibacter...     

Detection of ‘Candidatus Liberibacter asiaticus’ in citrus by concurrent tissue print-based qPCR and immunoassay

‘Candidatus Liberibacter asiaticus’ (CLas) is associated with the most destructive disease of citrus, huanglongbing (HLB). The most widely used methods for detection of CLas are PCR-based and require purification of DNA from plant samples. Elution of DNA from tissue prints made on nitrocellulose membranes followed by qPCR (TPE-qPCR) was compared to DNA extraction of plant tissue followed by PCR (X-PCR) by testing the same tissue samples. The former estimated a higher CLas population in tissue prints than the latter (t-test; P = 0.009). All extracts prepared for TPE-qPCR throughout the experiment were also tested by conventional PCR and 80.8% were identified as positive. A similar set of stem and petiole tissue samples was tested by TPE-qPCR and immunoassay. Although the detection rate by TPE-qPCR was higher than by immunoassay, about 6% of tissue prints were positive by immunoassay but not by TPE-qPCR. Thus, a higher detection rate would be achieved by combining TPE-qPCR with immunoassay. Significant differences were observed in the performance of nitrocellulose membranes from different manufacturers in these assays. Immunotissue prints showed that the spatial distribution pattern of CLas infection varied widely from one sample to another, but the patterns were highly correlated among serial sections from the same sample, suggesting that CLas preferentially colonizes adjacent phloem cells in a vertical rather than horizontal direction.     

Evidence for occurrence of an organophosphate-resistant type of acetylcholinesterase in strains of sea lice (Lepeophtheirus salmonis Krøyer)

Acetylcholinesterase (AChE) is the target of a major pesticide family, the organophosphates, which were extensively used as control agents of sea lice on farmed salmonids in the early 1990s. From the mid‐1990s the organophosphates dichlorvos and azamethiphos were seriously compromised by the development of resistance. AChE insensitive to organophosphate chemotherapeutants has been identified as a major resistance mechanism in numerous arthropod species, and in this study, target‐site resistance was confirmed in the crustacean Lepeophtheirus salmonis Krøyer isolated from several fish‐farming areas in Norway and Canada. A bimolecular rate assay demonstrated the presence of two AChE enzymes with different sensitivities towards azamethiphos, one that was rapidly inactivated and one that was very slowly inactivated. To our knowledge this is the first report of target‐site resistance towards organophosphates in a third class of arthropods, the Crustacea. Copyright © 2004 Society of Chemical Industry     

Impact of push–pull cropping system on pest management and occurrence of ear rots and mycotoxin contamination of maize in western Kenya

Push–pull involves intercropping of cereals with Desmodium as a “push” crop and planting Napier grass/Brachiaria as the “pull” crop at the border. The technology has been reported to effectively control stemborers, striga weed, and fall armyworm (FAW), and to improve soil nutrition, resulting in increased grain yield. This study evaluated the impact of stemborer and FAW management using this technology on incidence of maize ear rots and preharvest contamination of grains with aflatoxin and fumonisin in western Kenya. The study was conducted during three cropping seasons on maize grown under the push–pull system and as a monocrop. Incidence of stemborer and FAW damage was determined as percentage of damaged plants, while incidence of ear rots was determined as percentage of ears with symptoms. At harvest, fungi were isolated from kernels and aflatoxin and fumonisin were quantified using enzyme-linked immunosorbent assay. Stemborer and FAW damage was significantly (p = .001) reduced by over 50% under the push–pull system. There was also a significant (p < .001) reduction in the incidence of Fusarium verticillioides (60%) and Aspergillus flavus (86%), which was reflected in a reduced incidence of ear rots (50%) with the push–pull system (p = .001). Fumonisin in maize from push-pull farms was significantly (p = .048) reduced (39%) but the technology had no significant (p > .05) effect on aflatoxin. The study showed that push–pull is an effective strategy for managing maize ear rots and fumonisins, and therefore could play a role in improving food safety among smallholder maize farmers in the region.     

Tuber transmission of ‘Candidatus Liberibacter solanacearum’ and its association with zebra chip on potato in New Zealand

Zebra chip, an emerging disease of potatoes, has recently been associated with ‘Candidatus Liberibacter solanacearum’ in New Zealand. The phloem-limited bacterium is known to be vectored by the tomato potato psyllid (Bactericera cockerelli). In this study, the role of tuber transmission in the spread of Ca. L. solanacearum was investigated by re-planting potato tubers infected with Ca. L. solanacearum in the absence of the psyllid. Nested PCR demonstrated that Ca. L. solanacearum could be transmitted from the mother tubers both to the foliage of growing plants and to progeny tubers, resulting in symptomatic and asymptomatic plants. Of 62 Ca. L. solanacearum-infected tubers four did not sprout symptomatic of zebra chip. A further two plants developed foliar symptoms associated with zebra chip during the growing season and died prematurely. Fifty-six of the infected tubers produced asymptomatic plants, although Ca. L. solanacearum was detected in the foliage of 39 of them indicative of transmission into asymptomatic progeny plants. At harvest, Ca. L. solanacearum was found in the daughter tubers of only five of the 39 asymptomatic plants, and only one of these plants was found to have zebra chip symptoms in the daughter tubers. Our results show that tuber transmission of Ca. L. solanacearum could play a role in the life cycle of this pathogen, providing a source for acquisition by Bactericera cockerelli and for movement of the pathogen to other regions of New Zealand via transport of seed tubers.     

Scientific critique of the paper “Climatic distribution of citrus black spot caused by <Emphasis Type="Italic">Phyllosticta citricarpa</Emphasis>. A historical analysis of disease spread in South Africa” by Martínez-Minaya et al. (2015)

The global distribution of citrus black spot (CBS) disease, caused by Phyllosticta citricarpa, is climatically constrained, which is evident from its occurrence in citrus growing areas with warm, summer rainfall and its absence from areas with cooler, Mediterranean-type winter rainfall. Various epidemiological and modelling studies have supported this observation, predominantly estimating unsuitability for P. citricarpa in Mediterranean type climates, with no more than marginal suitability estimated at a few localities within some regions with Mediterranean type climates. The study by Martínez-Minaya et al. (European Journal of Plant Pathology, 143, 69–83, 2015), describes an historic sequence of recorded CBS occurrence in parts of South Africa, conducts an autocorrelation analysis and a correlative analysis with Köppen-Geiger climate zones and makes observations about the occurrence of certain Köppen-Geiger climate zones in the European Union. The study suggests that significant portions of the European Union and the broader Mediterranean basin are climatically similar to warm, summer rainfall areas in South Africa where P. citricarpa persists and causes CBS disease and concludes that the potential distribution of P. citricarpa is less constrained by climatic factors than spatial contagion. However, in this critique we expose methodological shortcomings in the Martínez-Minaya et al. (European Journal of Plant Pathology, 143, 69–83, 2015) study and conclude that the study grossly overestimated the extent of the geographical area that could support P. citricarpa, thereby rendering the findings scientifically unreliable.     

柑橘黄龙病症状与“Candidatus Liberibacter asiaticus”PCR检测结果的相关性分析

柑橘黄龙病症状较为复杂,且因寄主品种、生长期、病程等因素而异。利用PCR检测其病原菌"Candidatus Liberibacter spp."是目前柑橘黄龙病诊断的可靠方法之一。分析柑橘黄龙病症状与PCR检测结果的相关性有助于提高黄龙病的田间诊断准确率。本研究结果表明,与PCR检测相比,根据症状诊断柑橘黄龙病具有较高的假阳性率(8.20%)和假阴性率(50%);通过分析1 839个样品的症状与病原PCR检测结果发现,表现为斑驳型黄化、均匀黄化和"绿岛"这3种叶部症状以及"红鼻子果"和畸形果这2种果实症状的PCR病原检出率高;具有斑驳和黄化、黄化和"绿岛"、"绿岛"和花叶等复合症状样品的PCR检测"Ca.L.asiaticus"的阳性率最高;直径小于1 cm的幼果中的"Ca.L.asiaticus"检测稳定性低。这些结果为更准确地通过症状诊断柑橘黄龙病提供了科学依据。     

Numerical simulation of atmospheric transport of Phakopsora pachyrhizi urediniospores in South America using the state of Paraná-Brazil as a model

European Journal of Plant Pathology - Phakopsora pachyrhizi is a biotrophic fungus that needs living plant tissue in order to survive. The fungus causes Asian rust on soybean and costs billions of...     

The spread of the fall armyworm Spodoptera frugiperda in Africa—What should be done next?

Journal of Plant Diseases and Protection - The following text summarizes the different perspectives of presenters participating in the section Plant Protection in the Tropics and Subtropics, 61st...     

Rapid and quantitative detection of citrus huanglongbing bacterium ‘Candidatus Liberibacter asiaticus’ by real-time fluorescent loop-mediated isothermal amplification assay in China

Huanglongbing (HLB) is a devastating citrus disease that is associated with bacteria of the species ‘Candidatus Liberibacter asiaticus (Las)’ in China. All currently popular cultivars are susceptible to HLB, and there are no effective approaches to control the disease once the plants have been infected. Therefore, rapid and easy diagnostic tools are needed for maintaining clean nursery stock and inoculum removal strategies to control HLB. A real-time fluorescence loop-mediated isothermal amplification (RealAmp) targeting16S rDNA was developed for the rapid and quantitative detection of Las in China. The detection sensitivity of the RealAmp assay was approximately 1 pg/μl template DNA (equal to 2.4 × 10⁵ target DNA copies/μl) and no cross-reaction was observed with other pathogens or virus-free seedlings. Besides the quantitative detection using SYTO-9 fluorescent dye, an improved closed-tube visual inspection technique using SYBR Green I staining was developed for rapid screening of samples and minimize the risk of cross-contamination. The RealAmp assay is an alternative quantitative method, which would be used as a routine detection service for nursery materials in China.