Joseph E. Munyaneza

Association of Bactericera cockerelli (Homoptera: Psyllidae) with Zebra chip, a new potato disease in southwestern United States and Mexico

Abstract A new defect of potato,Solanum tuberosum L., “zebra chip,” so named for the characteristic symptoms that develop in fried chips from infected potato tubers, has recently been documented in several southwestern states of the United States, in Mexico, and in Central America. This defect is causing millions of dollars in losses to both potato producers and processors. Zebra chip plant symptoms resemble those caused by potato purple top and psyllid yellows diseases. Experiments were conducted to elucidate the association between the psyllidBactericera cockerelli (Sulc) (Homoptera: Psyllidae) and zebra chip by exposing clean potato plants to this insect under greenhouse and field conditions. Potato plants and tubers exhibiting zebra chip symptoms were tested for phytoplasmas by polymerase chain reaction. Potato psyllids collected from infected potato fields also were tested. Results indicated that there was an association between the potato psyllid and zebra chip. Plants exposed to psyllids in the greenhouse and field developed zebra chip. In the greenhouse, 25.8 and 59.2% of tubers exhibited zebra chip symptoms in the raw tubers and fried chips, respectively. In the field, 15 and 57% of tubers showed symptoms in raw tubers and chips, respectively. No zebra chip was observed in tubers from plants that had not been exposed to psyllids, either in the greenhouse or field. No phytoplasmas were detected from potato plants or tubers with zebra chip symptoms, suggesting that these pathogens are not involved in zebra chip. Of the 47 samples of potato psyllids tested, only two tested positive for the Columbia Basin potato purple top phytoplasma.

Zebra Chip Disease of Potato: Biology, Epidemiology, and Management

Zebra chip (ZC), a new and economically important disease of potato (Solanum tuberosum L.), has been documented to occur in commercial potato fields in the United States, Mexico, Central America, and New Zealand. This disease has caused millions of dollars in losses to the potato industry. Whole crops might be rejected because of ZC, often leading to abandonment of entire fields. Plant growth and yield are severely affected by the disease. Additionally, chips or fries processed from ZC-infected tubers exhibit dark stripes that become markedly more visible with frying, and hence are commercially unacceptable. The disease causes serious losses to the fresh market, tablestock and export potato industry as well. ZC-infected tubers usually do not sprout and if they do, produce hair sprouts or weak plants. Finally, there are indications that ZC symptoms might develop in tubers during storage. ZC has been associated with a previously undescribed species of liberibacter, tentatively named “Candidatus Liberibacter solanacearum”, also known as “Ca. L. psyllaurous”. The bacterium is transmitted to potato by the potato psyllid, Bactericera cockerelli (Šulc). All commercial potato cultivars appear to be susceptible to ZC, and management tactics targeted against the potato psyllid are currently the only means to effectively manage the disease. Furthermore, there are concerns about quarantine and trade issues in psyllid-affected regions because some countries may require that shipments of potatoes from certain growing regions be tested for the disease before the shipments are allowed entry. ZC history, geographic distribution, biology, epidemiology, and management are discussed herein.ResumenSe ha documentado que Zebra chip (ZC), una enfermedad nueva y económicamente importante de la papa (Solanum tuberosum L.), se presenta en los campos comerciales de papa en los Estados Unidos, México, América Central y Nueva Zelanda. Esta enfermedad ha causado millones de dólares en pérdidas a la industria de la papa. Se pudieran rechazar cultivos completos debido a ZC, que conducen a menudo al abandono total de los campos. Se afecta severamente el crecimiento y el rendimiento de la planta por la enfermedad. Además, las hojuelas o papas fritas procesadas de tubérculos infectados con ZC exhiben líneas oscuras que se vuelven marcadamente más visibles con el freído, y por lo tanto son inaceptables comercialmente. La enfermedad causa pérdidas serias al mercado fresco, al domestico y a la industria de la exportación también. Los tubérculos infectados con ZC generalmente no brotan, y si lo hacen, producen brotes ahilados y plantas débiles. Finalmente, hay indicaciones de que los síntomas de ZC pueden desarrollarse en los tubérculos durante el almacenamiento. Se ha asociado a la ZC con una especie previamente descrita de Liberibacter, tentativamente llamada “Candidatus Liberibacter solanacearum”, también conocida como “Ca. L. psyllaurous”. La bacteria se transmite a la papa por el psilido Bactericera cockerelli (Šulc). Todas las variedades comerciales de papa parecen ser susceptibles a ZC, y las tácticas de manejo dirigidas contra el psilido de la papa son a la fecha los únicos medios para manejar la enfermedad efectivamente. Aún mas, hay preocupación acerca de cuarentena y temas de comercio en regiones afectadas por el psilido, debido a que algunos países pudieran requerir que los embarques de papas de ciertas regiones de cultivo fueran analizados para la enfermedad antes de permitir la entrada de los envíos. De aquí que se discute la historia de ZC, la distribución geográfica, biología, epidemiología y el manejo.

Association of “Candidatus Liberibacter solanacearum” with the Psyllid, Trioza apicalis (Hemiptera: Triozidae) in Europe

ABSTRACT The psyllid Trioza apicalis Förster (Hemiptera: Triozidae) is a serious pest of carrots, Daucus carota L., in Europe. Carrots exhibiting symptoms of psyllid damage were observed in commercial fields in southern Finland in 2008. Symptoms in affected plants included leaf curling, yellow and purple discoloration of leaves, stunted growth of shoots and roots, and proliferation of secondary roots, Mechanisms by which T. apicalis induces symptoms in plants are not understood, and no plant pathogens have yet been associated with this insect, Given recent association of liberibacter with several crops affected by psyllids, an investigation on whether this bacterium is associated with T. apicalis was conducted, Polymerase chain reaction (PCR) primer pairs OA2/OI2c and LsoF/OI2c, specific for 16S rRNA gene from “Candidatus Liberibacter solanacearum,” generated amplicons of 1,168 bp and 1,173 bp, respectively, from DNA extracted from field-collected psyllids (61 and 36.6%, respectively), laboratory-reared psyllids (70 and 33.3%, respectively), field-collected petioles from symptomatic carrots (80 and 55%, respectively), and laboratory-grown carrots (100% for both primer pairs). In contrast, no PCR products were detected in DNA extracted from insect-free plants. The DNA sequences of amplicons of the genes encoding liberibacter 16S rRNA from psyllids and carrots were identical. DNA of the 16S rRNA gene sequences determined from carrots and psyllids were 99.9% identical to analogous sequences of “Ca. L. solanacearum” amplified from several solanaceous crops and the psyllid Bactericera cockerelli (Sulc), a vector of this bacterium, This is the first report of a plant pathogen associated with T. apicalis and the second known psyllid species associated with “Ca. L. solanacearum.”

Vector Transmission Efficiency of Liberibacter by Bactericera cockerelli (Hemiptera: Triozidae) in Zebra Chip Potato Disease: Effects of Psyllid Life Stage and Inoculation Access Period

ABSTRACT Successful transmission of plant pathogens by insects depends on the vector inoculation efficiency and how rapidly the insect can effectively transmit the pathogen to the host plant. The potato psyllid, Bactericera cockerelli (Šulc), has recently been found to transmit “Candidatus Liberibacter solanacearum,” a bacterium associated with zebra chip (ZC), an emerging and economically important disease of potato in several parts of the world. Currently, little is known about the epidemiology of ZC and its vectors inoculation capabilities. Studies were conducted in the field and laboratory to 1) assess transmission efficiency of potato psyllid nymphs and adults; 2) determine whether psyllid inoculation access period affects ZC incidence, severity, and potato yield; and 3) determine how fast the psyllid can transmit liberibacter to potato, leading to ZC development. Results showed that adult potato psyllids were highly efficient vectors of liberibacter that causes ZC and that nymphs were less efficient than adults at transmitting this bacterium. It was also determined that inoculation access period had little influence on overall ZC disease incidence, severity, and resulting yield loss. Moreover, results showed that exposure of a plant to 20 adult potato psyllids for a period as short as 1 h resulted in ZC symptom development. Furthermore, it was shown that a single adult potato psyllid was capable of inoculating liberibacter to potato within a period as short as 6 h, thereby inducing development of ZC. This information will help in developing effective management strategies for this serious potato disease.

High resolution melting analysis of the cytochrome oxidase I gene identifies three haplotypes of the potato psyllid in the United States.

ABSTRACT The potato psyllid, Bactericera cockerelli (Sulc) (Hemiptera: Triozidae), is a vector of the bacterium “Candidatus Liberibacter solanacearum,” the putative causal agent of potato zebra chip disease that has seriously affected the potato industry in the Central and Southwestern United States for the past decade. The 2011 potato growing season saw the first report of zebra chip disease in Washington, Oregon, and Idaho; however, B. cockerelli has been recorded in this region every season at least for the past 7 yr. Studies were conducted to determine the relationship between psyllids collected from the Pacific Northwest potatoes in 2011 and those from the Southwestern and Central United States. High resolution melting analysis of the B. cockerelli mitochondrial Cytochrome C Oxidase subunit I-like gene was conducted on over 450 psyllids collected from numerous locations across the Central and Western United States. Results suggest that at least three potato psyllid haplotypes exist in the United States, correlating to the Central, Western, and Northwestern United States geographical regions. The high resolution melting analysis results were subsequently supported by DNA sequencing data.

Haplotypes of "Candidatus Liberibacter solanacearum" suggest long-standing separation

Three haplotypes of the recently discovered bacterium species “Candidatus Liberibacter solanacearum” are described and related to geographic ranges. The first two are associated with Zebra Chip/Psyllid Yellows of potatoes and other solanaceous plants, vectored by the tomato/potato psyllid Bactericera cockerelli in North and Central America and New Zealand. The third is associated with diseased carrots in Finland and vectored by the carrot psyllid Trioza apicalis. The haplotypes are described by SNPs on the 16s rRNA, 16s/23s ISR and 50s rplJ and rplL ribosomal protein genes. These SNPs are inherited as a package across the three genes. Haplotype “a” has been found primarily from Honduras and Guatemala through western Mexico to Arizona and California, and in New Zealand. Haplotype “b” is currently known from eastern Mexico and northwards through Texas to south central Washington. These haplotypes show some range overlap in Texas, Kansas and Nebraska. The haplotypes are not yet known to elicit biological differences in the plant or insect hosts. These apparently stable haplotypes suggest separate bacterial populations of long standing.

Carrot Purple Leaf: A New Spiroplasmal Disease Associated with Carrots in Washington State

During the growing seasons of 2003 and 2004, a disease occurred in several carrot crops in south central Washington with symptoms suggestive of infection by phytopathogenic mollicutes (phytoplasmas and spiroplasmas). In the fall, many affected carrot plants exhibited extensive purple or yellow-purple leaf discoloration, general stunting of shoots and taproots, and formation of bunchy, fibrous secondary roots. For detection of the putative causal agents, polymerase chain reaction (PCR) assays were performed using primers specific to phytoplasmas as well as primers specific to plant-pathogenic spiroplasmas. Restriction fragment length polymorphism (RFLP) analyses of PCR-amplified 16S rDNA sequences revealed that about 81% of affected plants showing dark purple or yellow-purple leaf symptoms tested positive for Spiroplasma citri. Of affected plants showing mild purple discoloration of leaf margins, 18% tested positive for a phytoplasma strain belonging to the clover proliferation group (16SrVI), subgroup 16SrVI-A, and 11% for another phytoplasma strain belonging to the aster yellows group (16SrI), subgroup 16SrI-A. Nucleotide sequence analysis of cloned 16S rDNA confirmed the phytoplasma group affiliations. Some symptomatic plants were co-infected with S. citri and either aster yellows phytoplasma or clover proliferation group phytoplasma. To our knowledge, this is the first documentation of spiroplasma infection of carrot in the United States.

Psyllids as Vectors of Emerging Bacterial Diseases of Annual Crops

Abstract. Psyllids (Hemiptera: Psylloidea) are important pests of agricultural crops worldwide. These insects may cause damage to plants by direct feeding and/or vectoring plant pathogens. Psyllid-transmitted bacterial diseases are increasingly becoming important in perennial and annual crops. Several reports have shown that the fastidious bacterium ‘Candidatus Liberibacter’, vectored by at least four psyllid species, is associated with newly-emerging and economically important diseases of crops, including Huanglongbing or citrus greening disease in Citrus spp. and zebra chip in potatoes, Solanum tuberosum L. Huanglongbing is vectored by the Asian citrus psyllid, Diaphorina citri Kumayama, and the African citrus psyllid, Trioza erytreae Del Guercio, whereas zebra chip is vectored by the potato/tomato psyllid, Bactericera cockerelli Sulc Recently, ‘Ca. Liberibacter’ has been associated for the first time with the carrot psyllid, Trioza apicalis Förster, and carrot, Daucus carota L. subsp. sativus, plants affected by this insect in northern Europe. An overview of psyllid species vectoring bacterial diseases to annual crops, with emphasis on potato psyllid and carrot psyllid, is presented herein.

Detection of ‘Candidatus Liberibacter Solanacearum’ in the Potato Psyllid, Bactericera cockerelli (Sulc)1, by Conventional and Real-Time PCR

Abstract. ‘Candidatus Liberibacter solanacearum’ is the putative causal agent of zebra chip disease of potato, Solanum tuberosum L., and is transmitted by the potato psyllid, Bactericera cockerelli (Sulc). Much work has been done on the association of Liberibacter with the potato psyllid and several laboratories currently use polymerase chain reaction (PCR) to test for the bacterium. We have been testing for the bacterium in individual and composite samples of B. cockerelli as part of an ongoing investigation of the incidence of ‘Ca. L. solanacearum’ in potato psyllids collected in various parts of the US. Here we report that the bacterium can be reliably detected by using conventional and quantitative real-time PCR of nucleic acid extracts prepared from composite samples of one psyllid from a Liberibacterinfected colony combined with either one, two, three, four, five, nine, 14, 19, or 29 psyllids from a Liberibacter-free psyllid colony. This ability makes it possible to more easily test large numbers of psyllids for the presence of ‘Ca. Liberibacter solanacearum’. This, in turn, will aid efforts to predict the threat of developing serious zebra chip disease as a result of psyllid infestations in potato fields.

Phenotypic and Etiological Differences Between Psyllid Yellows and Zebra Chip Diseases of Potato

Both potato psyllid yellows and zebra chip (ZC) potato diseases are associated with the potato psyllid, Bactericera cockerelli (Sulc). Aboveground plant symptoms of both diseases are similar but there is a difference in symptoms in potato tubers. ZC has recently been associated with a new species of the bacterium liberibacter, ‘Candidatus Liberibacter solanacearum’, also known as ‘Ca. Liberibacter psyllaurous’. Mechanisms by which the potato psyllid might cause either ZC or potato psyllid yellows symptoms are not understood. Insect transmission studies were conducted to demonstrate psyllid vectoring of both diseases and to compare symptoms and development of the two diseases. Potato plants were exposed to both liberibacter-free and liberibacter-carrying potato psyllids and later evaluated for plant and tuber symptoms. These plants and tubers were then tested for liberibacter by polymerase chain reaction (PCR). In addition, potato plants exhibiting severe psyllid yellows/ZC-like symptoms were collected from a commercial potato field heavily infested with the potato psyllid and tested for liberibacter. PCR detected ‘Ca. Liberibacter solanacearum’ in ZC symptomatic plants and tubers resulting from exposure to liberibacter-carrying psyllids. Despite development of foliar symptoms that resemble those of ZC in plants exposed to liberibacter-free psyllids, no liberibacter was detected in these plants with psyllid yellows. Moreover, tubers from these plants with psyllid yellows did not exhibit any symptoms of ZC infection and tested negative for the bacterium. No liberibacter was detected in plants or tubers collected from the psyllid-infested potato field, suggesting that the observed symptoms were due to psyllid yellows. Furthermore, potato plants that were infected with liberibacter died sooner than plants that were infected with psyllid yellows. Although an association between liberibacter and ZC has been established, no pathogen is yet associated with potato psyllid yellows and mechanisms by which psyllid yellows symptoms are induced by the potato psyllid remain unclear.ResumenLas dos enfermedades de la papa, el amarillamiento de la papa por psílidos y zebra chip (ZC), están asociadas con el psílido de la papa Bactericera cockerelli (Sulc). Los síntomas aéreos de la planta por ambas enfermedades son similares, pero hay una diferencia en los síntomas del tubérculo. La ZC se ha asociado recientemente con una nueva especie de bacteria liberibacter, “Candidatus Liberibacter solanacearum” , también conocida como “Ca. Liberibacter psyllaurous”. No se han entendido los mecanismos por los cuales el psílido de la papa puede causar los síntomas, ya sea de la ZC o el amarillamiento de la papa por psílidos. Se condujeron estudios de la transmisión por insectos para demostrar la transmisión de ambas enfermedades por psílidos y para comparar los síntomas y el desarrollo de ambas enfermedades. Se expusieron plantas de papa a psílidos de la papa, tanto libres como con liberibacter, y se evaluaron posteriormente los síntomas de la planta y del tubérculo. Estas plantas y tubérculos se probaron después para liberibacter con la reacción en cadena de la polimerasa (PCR). Además, se colectaron plantas de papa que exhibieron síntomas severos de amarillamiento por psílidos/ZC, de un campo comercial de papa severamente infestado con el psílido de la papa, y se probaron para liberibacter. La PCR detectó “Ca. Liberibacter solanacearum” en plantas y tubérculos con síntomas de ZC, como resultado de su exposición a los psílidos con liberibacter. A pesar del desarrollo de síntomas foliares parecidos a los de ZC en plantas expuestas a los psílidos libres de liberibacter, no se detectó liberibacter en estas plantas con amarillamiento por psílidos. Aún mas, los tubérculos de estas plantas con amarillamiento por psílidos no exhibieron síntomas de infección por ZC y resultaron negativas a la bacteria. No se detectó liberibacter en plantas o tubérculos colectados del campo de papa infestado por psílidos, lo cual sugiere que los síntomas observados fueron debidos al amarillamiento por psílidos. Incluso, las plantas infectadas con liberibacter murieron más pronto que las infectadas con amarillamiento por psílidos. Aún cuando se ha establecido una asociación entre liberibacter y ZC, no se ha asociado a algún patógeno con el amarillamiento de la papa por psílidos y permanecen si aclararse los mecanismos por los cuales se inducen los síntomas del amarillamiento por el psílido de la papa.