Open AccessScreening nested-PCR primer for ‘Candidatus Liberibacter asiaticus’ associated with citrus Huanglongbing and application in Hunan, China
Author(s) -
Hong Yu,
Yoonsoo Hahn,
Jianglan Yi,
Ling He,
Liangying Dai,
Tuyong Yi
Publication year - 2019
Publication title -
plos one
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.99
H-Index - 332
ISSN - 1932-6203
DOI - 10.1371/journal.pone.0212020
Subject(s) - primer (cosmetics) , biology , nested polymerase chain reaction , candidatus , virology , microbiology and biotechnology , polymerase chain reaction , bacteria , 16s ribosomal rna , genetics , chemistry , gene , organic chemistry
Citrus Huanglongbing (HLB) is one of the most devastating citrus diseases worldwide. Sensitive and accurate assays are vital for efficient prevention of the spread of HLB-associated “ Candidatus Liberibacter spp ”. “ Candidatus Liberibacter spp ” that infect Citrus includes “ Candidatus Liberibacter asiaticus ” (Las), “ Candidatus Liberibacter africanus ” (Laf) and “ Candidatus Liberibacter americanus ” (Lam). Of them, Las is the most widespread species. In this study, a set of nested PCR primer pairs were screened to diagnose Las, and the nested PCR method greatly enhanced the sensitivity to detect Las up to 10 times and 100 times compared to qPCR and conventional PCR, respectively. Totally, 1112 samples from 5 different citrus cultivars in 39 different counties and cities were assayed by nested PCR. The results show that 384 samples were HLB-infected; the highest positive detection rate was 79.7% from the lopsided fruit samples, and the lowest positive detection rate was 16.3% from the apical dieback samples. The results indicate that the designed nested PCR primer pairs can detect Las from different symptomatic tissues, different citrus cultivars and different geographic regions. The set of nested PCR primers designed in the present study will provide a very useful supplementation to the current approaches for Las detection.