Survey for citrus greening disease in southern Iran

Huanglongbing (HLB), also known as greening, is a destructive disease of citrus that may limit production of this crop in many parts of Asia, Africa and South America. HLB is caused by a non-culturable phloem-limited bacterium of the genus 'Candidatus Liberibacter'. Three Candidatus species of Liberibacter, namely, 'Ca. L. asiaticus', 'Ca. L. africanus' and 'Ca. L. americanus', have been identified . The psyllid Diaphorina citri, vector of 'Ca. L. asiaticus' was first detected in December 1997 in Iran in an area close to the border with Pakistan. Since then, high populations of D. citri have been found in citrus plantations of Hormozgan and Kerman provinces in southern Iran. Based on the presence of psyllid D. citri, vector of 'Ca. L. asiaticus' and exibition of typical HLB symptoms, including mottling of leaves and yellowing of shoots in citrus growing areas of Sistan _ Baluchistan and Hormozgan provinces, samples of psyllids and symptomatic citrus trees were tested for the presence of HLB using biological indexing and PCR assay. D. citri was found in all citrus growing areas of Sistan- Baluchistan, Hormozgan and Kerman( Jiroft and Kahnooj) provinces. Nested PCR using primer pairs F1/R1 and F2/R2, specific for asiatic form of HLB, resulted in amplification of a 400 bp product from 29 symptomatic citrus plants( 28 Valencia sweet orange and a mandarin variety ) and 103 psyllid samples collected from Sistan _ Baluchistan and Hormozgan provinces. By direct PCR using Primer pair A2/J5 and nested PCR using Primer pairs OI1/OI2c and CGO3/CGO5, asiatic form of HLB was detected from 14 symptomatic Valencia and 2 local variety of sweet orang trees in Sistan _ Baluchistan province. No amplicons were obtained from symptomless sweet orange trees and psyllid samples reared on healthy sweet orange seedlings. The amplified fragment from a psyllid sample was cloned and sequenced (GenBank Accession No. FJ172759). BLAST search showed 100% identity with corresponding sequences of 'Ca. L. asiaticus' (M9439, AY34200 and EU078703). This is the first report on the occurrence of HLB disease in Iran. The disease appears to be widely distributed in citrus growing regions of southern Iran and may be a component of citrus decline in this area. To prevent further spread of HLB, quarantine measures must be implemented and inoculum eliminated by removal of infected trees and chemical treatment against vector.