Plant Science

The Asian citrus psyllid, Diaphorina citri, is an invasive insect and a vector of ‘ Candidatus Liberibacter asiaticus’ ( CLas), a bacterium whose growth in Citrus species results in huanglongbing (HLB), also known as citrus greening disease. Methods to enrich and sequence CLas from D. citri often rely on biased genome amplification and nevertheless contain significant quantities of host DNA. To overcome these hurdles, we developed a simple pretreatment DNase and filtration (PDF) protocol to remove host DNA and directly sequence CLas and the complete, primarily uncultivable microbiome from D. citri adults. The PDF protocol yielded CLas abundances upward of 60% and facilitated direct measurement of CLas and endosymbiont replication rates in psyllids. The PDF protocol confirmed our lab strains derived from a progenitor Florida CLas strain and accumulated 156 genetic variants, underscoring the utility of this method for bacterial strain tracking. CLas genetic polymorphisms arising in lab-reared psyllid populations included prophage-encoding regions with key functions in CLas pathogenesis, putative antibiotic resistance loci, and a single secreted effector. These variants suggest that laboratory propagation of CLas could result in different phenotypic trajectories among laboratories and could confound CLas physiology or therapeutic design and evaluation if these differences remain undocumented. Finally, we obtained genetic signatures affiliated with Citrus nuclear and organellar genomes, entomopathogenic fungal mitochondria, and commensal bacteria from laboratory-reared and field-collected D. citri adults. Hence, the PDF protocol can directly inform agricultural management strategies related to bacterial strain tracking, insect microbiome surveillance, and antibiotic resistance screening.

Citrus Huanglongbing (HLB), known as the most economically devastating disease in citrus industry, is mainly caused by phloem-restricted Gram-negative bacterium “Candidatus Liberibacter asiaticus” (CLas). To date, CLas is still unculturable in vitro, which has been dramatically delaying the research on its pathogenesis, and only few Sec-dependent effectors (SDEs) have been identified to elucidate the pathogenesis of CLas. Here, we confirmed that a CLas-secreted Sec-dependent polypeptide, namely SECP8 (CLIBASIA_05330), localized in nucleus, cytoplasm and cytoplasmic membrane, and showed remarkably higher transcript abundance in citrus than in psyllids. Potato virus X (PVX)-mediated transient expression assays indicated that mSECP8 (the mature form of SECP8) suppressed pro-apoptotic mouse protein BAX and Phytophthora infestans elicitin INF1-triggered hypersensitive response (HR) associated phenotypes, including cell death, H2O2 accumulation and callose deposition. Intriguingly, mSECP8 also inhibited SDE1 (CLIBASIA_05315)-induced water-soaked and dwarfing symptoms in Nicotiana benthamiana. In addition, mSECP8 can promote the susceptibility of transgenic Wanjincheng orange (Citrus sinensis) to CLas invasion and further HLB symptom development, and it contributes to the proliferation of Xanthomonas citri subsp. citri (Xcc). Moreover, the expression of ten immunity-related genes were significantly down-regulated in mSECP8 transgenic citrus than those in wide-type (WT) plants. Overall, we propose that mSECP8 may serve as a novel broad-spectrum suppressor of plant immunity, and provide the first evidence counteractive effect among CLas effectors. This study will enrich and provide new evidences for elucidating the pathogenic mechanisms of CLas in citrus host.

Background: Alfalfa, Medicago sativa L. is the most important and widely grown leguminous fodder crop in temperate and tropical regions of the world. The production of alfalfa crop is limited by several biotic stresses, among which witches’ broom disease (AWB) was reported to cause significant economic losses. Methods: The phytoplasma infected alfalfa plants were collected from a central research farm, ICAR-IGFRI, Jhansi, U.P. Qualitative parameters such as crude protein content, acid detergent fibre and neutral detergent fibre were estimated in diseased and healthy plants. Phytoplasma universal primer (P1/P7) and nested primer (R16mF2/R16mR1) were used for the molecular characterization of AWB infected plants and phytoplasma infected, Parthenium hysterophorus. Result: The incidence of AWB disease ranged from 8-10%. The quantitative analysis of disease plants showed reduced plant height (-35%), fresh weight (-46.89%) and dry weight (-50.08%) compared to healthy plants. The diseased plant recorded low crude protein content (-21.38%) and higher dry matter content (+0.68%), acid detergent fibre (+33.72%) and neutral detergent fibre (+13.06%). The association of phytoplasma in diseased alfalfa and parthenium samples was confirmed by using P1/P7 and R16mF2/R16mR1 primer pair and Blastn analysis shared 99.6-100% similarity with ‘Candidatus Phytoplasma australasia’ belongs to the 16Sr group II-D.

Las variedades de papa (Solanum tuberosum L.) producidas en México son susceptibles a Candidatus Liberibacter solanacearum (CaLso), causante de la enfermedad conocida como ‘manchado interno de la pulpa’, por lo que se requiere conocer la respuesta de genotipos experimentales a la bacteria. El presente estudio tuvo como objetivo evaluar el efecto de la infección de los haplotipos LsoA + LsoB de CaLso en el follaje, la biomasa seca y la calidad de tubérculo de papa, variedad Fianna, una colecta de Solanum demissum y los clones experimentales T90-1-63 y T05-13-21 de Solanum spp. El manchado interno de la pulpa del tubérculo se determinó mediante análisis de imágenes de tubérculos. Las plantas de Fianna mostraron la mayor severidad de daño foliar; en cambio, los clones experimentales presentaron 17 % menos daño foliar que Fianna y 8 % más daño foliar que S. demissum. Este último fue el genotipo con la mayor biomasa seca de hoja y produjo tubérculos de un tamaño pequeño; las plantas infectadas de S. demissum presentaron mayor número de tubérculos y mayor rendimiento de tubérculo fresco que las plantas sin inoculación, aunque tuvieron la menor proporción de superficie sana del tubérculo. El clon T90-1-63 presentó los porcentajes más altos de superficie sana de tubérculo (> 79 %) y las menores intensidades del manchado interno de la pulpa del tubérculo.