Identification of Autophagy-Related Genes in the Potato Psyllid, Bactericera cockerelli and Their Expression Profile in Response to ‘Candidatus Liberibacter Solanacearum’ in the Gut

Autophagy, also known as type II programmed cell death, is a cellular mechanism of “self-eating”. Autophagy plays an important role against pathogen infection in numerous organisms. Recently, it has been demonstrated that autophagy can be activated and even manipulated by plant viruses to facilitate their transmission within insect vectors. However, little is known about the role of autophagy in the interactions of insect vectors with plant bacterial pathogens. ‘Candidatus Liberibacter solanacearum’ (Lso) is a phloem-limited Gram-negative bacterium that infects crops worldwide. Two Lso haplotypes, LsoA and LsoB, are transmitted by the potato psyllid, Bactericera cockerelli and cause damaging diseases in solanaceous plants (e.g., zebra chip in potatoes). Both LsoA and LsoB are transmitted by the potato psyllid in a persistent circulative manner: they colonize and replicate within psyllid tissues. Following acquisition, the gut is the first organ Lso encounters and could be a barrier for transmission. In this study, we annotated autophagy-related genes (ATGs) from the potato psyllid transcriptome and evaluated their expression in response to Lso infection at the gut interface. In total, 19 ATGs belonging to 17 different families were identified. The comprehensive expression profile analysis revealed that the majority of the ATGs were regulated in the psyllid gut following the exposure or infection to each Lso haplotype, LsoA and LsoB, suggesting a potential role of autophagy in response to Lso at the psyllid gut interface.

Exogenous Application of Polycationic Nanobactericide on Tomato Plants Reduces the Candidatus Liberibacter Solanacearum Infection

Candidatus Liberibacter solanacearum (CaLso) is associated with diseases in tomato crops and transmitted by the tomato psyllid Bactericera cockerelli. A polymeric water-dispersible nanobactericide (PNB) was evaluated against CaLso as a different alternative. PNB is a well-defined polycationic diblock copolymer designed to permeate into the vascular system of plants. Its assessment under greenhouse conditions was carried out with tomato plants previously infected with CaLso. Using a concentration as low as 1.0 mg L−1, a small but significant reduction in the bacterial load was observed by real-time qPCR. Thus, to achieve an ecologically friendly dosage and set an optimum treatment protocol, we performed experiments to determine the effective concentration of PNB to reduce ~65% of the initial bacterial load. In a first bioassay, a 40- or 70-fold increase was used to reach that objective. At this concentration level, other bioassays were explored to determine the effect as a function of time. Surprisingly, a real reduction in the symptoms was observed after three weeks, and there was a significant decrease in the bacterial load level (~98%) compared to the untreated control plants. During this period, flowering and formation of tomato fruits were observed in plants treated with PNB.

Dual transcriptional profiling of carrot and ‘Candidatus Liberibacter solanacearum’ at different stages of infection suggests complex host-pathogen interaction

The interactions between the phloem-limited pathogen ‘Candidatus Liberibacter solanacearum’ (CLso) haplotype C and carrot (Daucus carota subsp. sativus) were studied at 4, 5 and 9 weeks post inoculation (wpi), by combining dual RNA-Seq results with data on bacterial colonization and observations of the plant phenotype. In the infected plants, genes involved in jasmonate biosynthesis, salicylate signaling, PAMP and effector-triggered immunity and production of pathogenesis-related proteins were upregulated. At 4 wpi, terpenoid synthesis-related genes were upregulated, presumably as a response to the psyllid feeding, whereas at 5 and 9 wpi, genes involved in both the terpenoid and flavonoid production were downregulated and phenylpropanoid genes were upregulated. Chloroplast-related gene expression was downregulated, in concordance with the observed yellowing of the infected plant leaves. Both the RNA-Seq data and electron microscopy suggested callose accumulation in the infected phloem vessels, likely to impair the transport of photosynthates, while phloem regeneration was suggested by the formation of new sieve cells and the upregulation of cell wall-related gene expression. The CLso genes involved in replication, transcription and translation were expressed at high levels at 4 and 5 wpi, whereas at 9 wpi the Flp pilus genes were highly expressed, suggesting adherence and reduced mobility of the bacteria. The CLso genes encoding ATP and C4-dicarboxylate uptake were differentially expressed between the early and late infection stages, suggesting a change in the dependence on different host-derived energy sources. HPE1 effector and salicylate hydroxylase were expressed, presumably to suppress host cell death and salicylic acid-dependent defenses during the infection.

Effect of Plant Age, Temperature and Vector Load on ‘Candidatus Liberibacter solanacearum’ in planta titer and Shoot Proliferation Symptoms in Carrot

A decade ago, shoot proliferation symptoms (witches’ broom) in carrots were believed to be the cause of ‘Candidatus Phytoplasma’ and/or Spiroplasma infection, yet in recent years, this association appeared to have weakened and a closer association was found with the yet-unculturable, psyllid-transmitted Gram-negative bacterium, ‘Candidatus Liberibacter solanacearum’. In Israel, carrots are grown throughout the year, yet shoot proliferation symptoms tend to appear only in mature plants and mostly during late spring to early summer. We hypothesized that factors such as plant age, temperature and vector load, which vary along the year, have a critical effect on symptoms development and set to examine these factors under controlled conditions. Here we show that young carrot seedlings are as prone as older plants, to develop shoot proliferation symptoms, following ‘Ca. L. solanacearum’ inoculation. Surprisingly, we found that the local ‘Ca. L. solanacearum’ haplotype was extremely sensitive to constant temperature of 30˚C, which led to a significant reduction in bacterial growth and symptoms development, compared with 18˚C which was very conducive for symptoms development. We have also found that inoculations with 10 or 20 psyllids per plant results in faster symptoms development compared with inoculations with 2 psyllids per plant, however, the disease progress rate was insignificant among the different vector loads. These data provide important insight to the effects of plant age, temperature and vector load on ‘Ca. L. solanacearum’ and its associated symptoms and strengthen the notion that ‘Ca. L. solanacearum’ is the main responsible agent for carrot witches broom in Israel.

Bactericera cockerelli vector de Candidatus Liberibacter solanacearum, morfometría y haplotipos en poblaciones de México

Bactericera cockerelli es una plaga de importancia económica en solanáceas en México, por los amarillamientos que causa en los cultivos, así como por la transmisión de Candidatus Liberibacter solanacearum. Se describen variantes genéticas de este insecto, las cuales se relacionan con su capacidad para fungir como vector. En México, la distribución de B. cockerelli es muy amplia y se carece de información acerca de sus características morfológicas y genéticas. El objetivo de esta investigación fue caracterizar morfológica y genéticamente a B. cockerelli y detectar la presencia de Ca. L. solanacearum en poblaciones de B. cockerelli de las zonas productoras de solanáceas en México. Para lo cual se muestrearon 35 localidades de 13 estados, sobre cultivos de chile, tomate, berenjena y papa, bajo diferentes sistemas de producción. Se midieron las variables largas de cuerpo (LC), largas de abdomen (LAB) y ancho de abdomen (AAB) en insectos de cada población, se detectó la presencia de Ca. L. solanacearum en los 13 estados muestreados, donde los machos presentaron el mayor porcentaje de insectos positivos. La presencia de Ca. L. solanacearum no se vio influenciada por el hospedero o el sistema de producción, sino por la presencia de B. cockerelli.

Bactericera cockerelli Sulc. (Hemíptera: Triozidae) causante de punta morada (Candidatus liberibacter, solanacearum) en papa (Solanum tuberosum L.) en Estelí, Nicaragua

La papa Solanum tuberosum L pertenece a la familia de las solanáceas, es el cuarto cultivo sembrado en más de 100 países incluyendo Nicaragua, en este país existe un consumo por persona de hasta unos 8 kg anuales, la papa se cultiva en Nicaragua entre 800 a 1 200 hectáreas y se obtiene una producción que representa de 35 % a 40 % de la demanda nacional. El objetivo del presente estudio fue determinar la fluctuación poblacional de Bactericera cockerelli e incidencia de punta morada (Candidatus liberibacter, Solanacearum) en lotes comerciales de papa en el municipio de Estelí entre los meses de enero a noviembre de 2014, la toma de datos se realizó en lotes de papa en las localidades de Miraflor y El Tisey, los lotes fueron seleccionados con similitud en condiciones climáticas y de manejo del cultivo, en cada lote seleccionado se colocaron trampas amarillas con pegamento para la captura de adultos de B. cockerelli, de igual manera se realizaron muestreos aleatorios en 100 plantas por lote para medir la fluctuación poblacional de ninfas y adultos del vector, así como para medir la incidencia de síntomas similares a Candidatus liberibacter, solanacearum de las cuales se seleccionaron cinco plantas para análisis y detección del patógeno a través de la técnica de PCR en el Centro Nacional de Diagnostico Fitosanitario del IPSA. Los resultados indican que el insecto estuvo presente desde el inicio del estudio siendo las mayores capturas en los meses febrero a mayo; julio, agosto y octubre con 0.84 insectos promedio por trampa, de igual manera los meses de marzo, abril, julio y octubre fueron donde se expresaron más plantas con síntomas de 3 al 4% incidencia, así mismo de las muestras tomadas solamente el 20% resultó positivas a la enfermedad coincidiendo con baja poblaciones del insecto vector.

Candidatus Liberibacter solanacearum (zebra chip).

Candidatus Liberibacter solanacearum (Lso) is a phloem-limited, Gram-negative, unculturable bacterium that is primarily spread by psyllid insect vectors. It is considered very invasive due to its ability to be transported primarily in infective psyllids (Munyaneza et al., 2007a; 2010a,b; 2012a,b; Munyaneza, 2012; Alfaro-Fernandez et al., 2012a,b). It has been shown that Lso distribution in the Americas, New Zealand and Europe follows the distribution of its known psyllid vectors (Munyaneza, 2010; 2012).In New Zealand, where Lso was introduced along with Bactericera cockerelli, supposedly from Western USA in early 2000s, the bacterium had already spread to both North and South Island by the time it was first documented in 2006 (Gill, 2006). It is clear that introduction of the psyllid vectors of Lso into new regions is likely to result in the rapid spread of this bacterium. Lso and several of its vectors are already on several alert lists, including the EPPO A1 Regulated Quarantine Plant Pests.

Parámetros agronómicos del tomate (Solanum lycopersicum L.) inoculado con “Candidatus Liberibacter solanacearum” y tratados con fosfitos

La enfermedad del permanente del tomate o zebra chip cau- sada por “Candidatus Liberibacter solanacearum” (Ca. L. solanacearum), es de gran importancia en las regiones productoras de tomate en México, la cual causa pérdidas de entre el 60 y 100% de la producción. El objetivo del trabajo fue evaluar el efecto de seis fosfitos (FOS), ácido salicílico y acibenzolar s-metil sobre el crecimiento del cultivo de tomate y la severidad de Ca. L. solanacearum. Las plantas de tomate se infectaron con Ca. L. solanacearum mediante el confinamiento con adultos de B. cockerelli portadores de esta bacteria. Como respuesta se evaluaron la disminución de la severidad de la enfermedad y nueve parámetros agronómicos. La aspersión de fosfitos redujo la severidad de los síntomas de la enfermedad ocasionada por Ca. L. solanacearum, mejorando las variables altura de planta, peso del fruto, peso de planta y peso de raíz. En cambio, la aspersión de ácido salicílico y acibenzolar s-metil no redujeron los efectos de la enfermedad en el cultivo de tomate.