ABSTRACT
Aims: To identify and characterize Citrus Huanglongbing disease causing pathogen Candidatus Liberibacter asiaticus and to evaluate its biological control using medicinal plant extracts. Study Design: The study was designed based on standard laboratory protocol. Place and Duration of Study: Professor Joarder DNA and Chromosome Research Lab, Department of Genetic Engineering and Biotechnology, University of Rajshahi, Rajshahi-6205, Bangladesh between June 2015 and September 2016. Methodology: Causal pathogen of Citrus Huanglongbing disease was isolated from infected leaves. Different types of biochemical and morphological characterizations of Candidatus Liberibacter asiaticus were done. 16S rDNA primers (27F and 1391R) were used to amplify genomic DNA of Candidatus Liberibacter asiaticus. Sequencing of 16S rDNA sequence of Candidatus Liberibacter asiaticus were performed. Sensitivity pattern of Candidatus Liberibacter asiaticus against several standard antibiotics were done. Antimicrobial activity test was observed using two solvents extracts of four medicinal plants by disc diffusion method in vitro condition. Results: Candidatus Liberibacter asiaticus showed positive and negative response to different biochemical test. Candidatus Liberibacter asiaticus showed gram negative in gram staining test. Candidatus Liberibacter asiaticus showed highest 35±0.5 mm and lowest 8±0.2 mm zone of inhibition against amoxycillin and kanamycin respectively. Approximately 1300bp band was found in PCR amplification and phylogenetic tree analysis of 16S rRNA gene sequence of Candidatus Liberibacter asiaticus showed 75% similarity with Candidatus Liberibacter asiaticus strain 374.15. Candidatus Liberibacter asiaticus showed highest 16±0.5 mm diameter of zone of inhibition at 60 µg/ml concentration for ethanol extract of Cuscuta reflexa. Conclusion: This study will be helpful for proper identification, characterization and control of Citrus Huanglongbing disease causing pathogen Candidatus Liberibacter asiaticus in an eco-friendly way.
ABSTRACT
Citrus Huanglongbing (HLB) also known as citrus greening is one of the most devastating diseases of citrus worldwide. The disease is caused by Candidatus Liberibacter bacterium, vectored by the psyllid Diaphorina citri Kuwayama and Trioza erytreae Del Guercio. Citrus plants infected by the HLB bacterium may not show visible symptoms sometimes for years following infection. The aim of this study was to develop effective gene-specific primer pairs for polymerase chain reaction based method for quick screening of HLB disease. Thirty-two different gene-specific primer pairs, across the Ca. Liberibacter asiaticus genome, were successfully developed. The possibility of these primer pairs for cross-genome amplification across ‘Ca. Liberibacter africanus’ and ‘Ca. Liberibacter americanus’ were tested. The applicability of these primer pairs for detection and differentiation of Ca Liberibacter spp. is discussed.
ABSTRACT
En México y Centro América se han detectado tubérculos de papa con manchado interno. Recientemente en Texas EUA a esta enfermedad se le ha denominado "Zebra Chip" (ZC) o rayado de la papa, los síntomas foliares se asemejan al síndrome denominado "Punta Morada de la Papa" (PMP) o enfermedad del "amarillamiento por psilidos" la cual es asociada con la presencia de "Candidatus Liberibacter solanacearum". El objetivo de esta investigación fue detectar la presencia de esta bacteria y de fitoplasmas en plantas de papa que presentaban la coloración purpura de los foliolos. Durante el ciclo primavera - verano 2011 y 2012 se hizo un muestreo en los municipios de Tenango del Valle, Zinacantepec, Villa de Allende y San José del Rincón, del Estado de México. La detección de ambos patógenos se realizó mediante la reacción en cadena de la polimerasa (PCR) con los iniciadores específicos para fitoplasmas: P1/P7, R16mF2/R16mR1 y para Ca Liberibacter solanacearum: OA2/Oi2c, resultando el 35,8% de las plantas positivas para fitoplasmas y el 11,6% para la bacteria. Los resultados indican que en algunas regiones productoras de papa del Estado de México, los dos presuntos agentes causales del síndrome de PMP, fitoplasmas y Ca. Liberibacter solanacearum, pueden estar asociados.
In Mexico and Central America have been detected stained potato tubers with internal browning; recently in Texas, USA, this disease has been called "Zebra Chip" (ZC) or striped potato, foliar symptoms resemble the syndrome called "Potato Purple Top" (PPT) or "psyllid yellows" disease which is associated with the presence of "Candidatus Liberibacter solanacearum". The aim of the current work was to detect the presence of this bacterium and phytoplasma in potato plants with purple top symptoms. During 2011 and 2012 Spring - Summer cycle, a directed sampling was carried out in Tenango del Valle, Zinacantepec, Villa de Allende and San José del Rincón, State of México. The detection of both pathogens was performed by Polymerase Chain Reaction (PCR) with specific primers for phytoplasmas: P1/ P7, R16mF2/R16mR1 and for Ca Liberacter solanacearum: OA2/Oi2c, being 35,8 % from the positive plants for phytoplasmas and 11,6 % for this bacterium. These results indicated that in some areas these two PPT syndrome suspected causative agents, phytoplasmas and Ca. Liberibacter solanacearum, could be associated in the State of Mexico potato-producing region.