Detection and Molecular Characterization of a 16SrII‐A* Phytoplasma in Grapefruit (Citrus paradisi) with Huanglongbing‐like Symptoms in China

Abstract

In the year 2010, in a survey in Guangxi Province, China, to detect and characterize phytoplasmas in a huanglongbing (HLB)-infected grapefruit (Citrus paradisi) orchard, 87 leaf samples with symptoms of blotchy mottle were collected from symptomatic grapefruit trees, and 320 leaf samples from symptomless trees adjacent to the symptomatic trees. Nested polymerase chain reaction (PCR) using universal phytoplasma primer set P1/ P7 followed by primer set fU5/rU3 identified 7 (8.0%) positive samples from symptomatic samples but none from symptomless samples. Of the 87 symptomatic samples, 77 (88.5%) were positive for 'Candidatus Liberibacter asiaticus' and 5 for both phytoplasma and 'Ca. L. asiaticus'. Sequence analysis indicated that seven 881-bp amplicons, amplified by nested phytoplasma primer sets P1/P7 and fU5/rU3, shared 100.0% sequence identity with each other. Genome walking was then performed based on the 881 bp known sequences, and 5111 bp of upstream and downstream sequences were obtained. The total 5992 bp sequences contained a complete rRNA operon, composed of a 16S rRNA gene, a tRNA Ile gene, a 23S rRNA gene and a 5S rRNA gene followed by eight tRNA genes. Phylogenetic analysis and virtual restriction fragment length polymorphism analysis confirmed the phytoplasma was a variant (16SrII-A*) of phytoplasma subgroup 16SrII-A. As phytoplasmas were only detected in blotchy-mottle leaves, the 16SrII-A* phytoplasma identified was related to HLB-like symptoms. J Phytopathol

First report of ‘Candidatus Phytoplasma trifolii’ (16SrVI group) in Nagpur mandarin (Citrus reticulata) showing huanglongbing symptoms in central India

“…Although the disease is primarily attributed to 'Candidatus Liberibacter asiaticus', an unculturable α-proteobacterium, phytoplasmas belonging to the groups 16SrIX in Brazil (Texeira et al, 2008), 16SrII in China (Lou et al, 2014), and 16SrI in China (Chen et al, 2009) Liberibacter asiaticus' was tested by real-time PCR (ABI 7300, Applied Biosystems, USA) using specific16S rDNA-based TaqMan primer-probe sets (HLBasr/HLBp) (Li et al, 2006). To ascertain the association of phytoplasma, PCR was performed using P1/P7 universal primers specific to the 16S rRNA gene of phytoplasmas, followed by a nested PCR from a 1:30 dilution of the P1/P7 PCR products using primer sets R16F2n/R16R2 (Gundersen & Lee, 1996) and Fu5/Ru3 (Lou et al, 2014).…”

Detection and Inoculation of Peanut Witches' Broom Phytoplasma (16SrII‐A) and Periwinkle Leaf Yellowing Phytoplasma (16SrI‐B) in Citrus Cultivars in Taiwan

“…; Lou et al. ). The phytoplasma related to the pigeon pea witches' broom (PPWB) phytoplasma (16Sr IX) was found in sweet orange trees closely associated with HLB symptoms in São Paulo, Brazil, in 2008.…”

“…() and Lou et al. () in China recently reported that phytoplasmas are associated with HLB symptoms. However, no phytoplasma was found in HLB‐diseased citrus trees and in psyllids collected from Florida (Meyer and Hoy ; Sagaram et al.…”

“…The Citrus blotchy‐mottle (CBM) phytoplasma belonging to 16SrII‐A was detected in grapefruit trees with HLB‐like symptoms as leaf blotchy mottling at a low ratio (7/87) in Guangxi, China (Lou et al. ). Among seven positive samples, five samples were co‐infected with Las.…”

Diversity and characterization of Candidatus Liberibacter asiaticus strains causing huanglongbing disease in Iran, based on two prophage loci

“…P. asteris (Chen et al., ) and Ca . P. aurantifolia (in China) (Lou et al., ), Ca . P. asteris (in Mexico) (Arratia‐Castro et al., ) and Ca .…”