title: Development of rapid, sensitive and non-radioactive tissue-blot diagnostic method for the detection of citrus greening.
abstract: Citrus huanglongbing (HLB or citrus greening) is one of the most devastating diseases of citrus worldwide. The disease is caused by Gram-negative, phloem-limited α-proteobacterium, 'Candidatus Liberibacter asiaticus', vectored by the psyllid, Diaphorina citri Kuwayama. Citrus plants infected by the HLB bacterium may not show visible symptoms sometimes for years following infection and non-uniform distribution within the tree makes the detection of the pathogen very difficult. Efficient management of HLB disease requires rapid and sensitive detection early in the infection followed by eradication of the source of pathogen and the vector. The polymerase chain reaction (PCR) based method is most commonly employed for screening the infected/suspected HLB plants and psyllids. This is time consuming, cumbersome and not practical for screening large number of samples in the field. To overcome this, we developed a simple, sensitive, non-radioactive, tissue-blot diagnostic method for early detection and screening of HLB disease. Digoxigenin labeled molecular probes specific to 'Ca. L. asiaticus' nucleotide sequences have been developed and used for the detection of the pathogen of the HLB disease. The copy number of the target genes was also assessed using real-time PCR experiments and the optimized real-time PCR protocol allowed positive 'Ca. L. asiaticus' detection in citrus samples infected with 'Ca. L. asiaticus' bacterium.
authors: University of Florida, IFAS, Citrus Research and Education Center, 700 Experiment Station Road, Lake Alfred, FL 33850, USA; Department of Plant Sciences, The University of Tennessee, 252 Ellington Plant Sciences, 2431 Joe Johnson Dr., Knoxville, TN 37996, USA. Electronic address: mnrbhav@yahoo.com.
authors: Madhugiri
authors: M
authors: Nageswara-Rao
authors: Shin-Ichi
authors: S
authors: Miyata
authors: Dilip
authors: D
authors: Ghosh
authors: Mike
authors: M
authors: Irey
authors: Stephen M
authors: SM
authors: Garnsey
authors: Siddarame
authors: S
authors: Gowda
mesh_terms: D000818:Animals; D002957:Citrus / Q000382:microbiology*; D004269:DNA, Bacterial / Q000032:analysis; D004076:Digoxigenin / Q000737:chemistry; D006430:Hemiptera / Q000382:microbiology; D007303:Insect Vectors / Q000382:microbiology; D015335:Molecular Probes; D010935:Plant Diseases / Q000382:microbiology*; D018515:Plant Leaves / Q000382:microbiology; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D012230:Rhizobiaceae / Q000235:genetics / Q000302:isolation & purification*; D012680:Sensitivity and Specificity
chemical_list: D004269:DNA, Bacterial; D015335:Molecular Probes; D004076:Digoxigenin
keywords: Citrus greening; DNA probe; Psyllids; Real-time-PCR; Tissue blotting