Hunan Provincial Key Laboratory for Biology and Control of Plant Pests, College of Plant Protection, Hunan Agricultural University, Changsha, Hunan province, China.
College of life Science and Technology, Beijing University of Chemical Technology, Beijing, China.
PLoS One. 2019 Feb 22;14(2):e0212020. doi: 10.1371/journal.pone.0212020. eCollection 2019.
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.
柑橘黄龙病（HLB）是世界上最具破坏性的柑橘病害之一。对于有效预防 HLB 相关“韧皮杆菌属”的传播，灵敏和准确的检测至关重要。感染柑橘的“韧皮杆菌属”包括“亚洲韧皮杆菌”（Las）、“非洲韧皮杆菌”（Laf）和“美洲韧皮杆菌”（Lam）。其中，Las 分布最广。在这项研究中，筛选了一组巢式 PCR 引物对来诊断 Las，与 qPCR 和常规 PCR 相比，巢式 PCR 方法将检测 Las 的灵敏度分别提高了 10 倍和 100 倍。总共用巢式 PCR 对来自 39 个不同县/市的 5 种不同柑橘品种的 1112 个样本进行了检测。结果表明，384 个样本受到 HLB 感染；来自畸形果实的阳性检出率最高，为 79.7%，来自梢端枯死样本的阳性检出率最低，为 16.3%。结果表明，设计的巢式 PCR 引物对可以从不同的症状组织、不同的柑橘品种和不同的地理区域检测到 Las。本研究设计的巢式 PCR 引物集将为 Las 的检测提供非常有用的补充。
