Development and Evaluation of a Nested PCR for Improved Diagnosis and Genetic Analysis of Peste des Petits Ruminants Virus (PPRV) for Future Use in Nascent PPR Eradication Programme.
Development and Evaluation of a Nested PCR for Improved Diagnosis and Genetic Analysis of Peste des Petits Ruminants Virus (PPRV) for Future Use in Nascent PPR Eradication Programme.
复制标题
DOI：
10.3390/ani11113170
复制
发表时间：
2021-11-05
期刊：
影响因子：
--
通讯作者：
Parida S
中科院分区：
文献类型：
--
作者：
Mahapatra M;Neto MM;Khunti A;Njeumi F;Parida S
小反刍动物疫病(PPR)是一种对小型反刍动物和大量野生动物具有高度传染性和重要经济价值的病毒性疾病。病原体PPR病毒(PPRV)主要在非洲、中东和亚洲的大片地区传播，幸运的是，目前正在接受全球根除计划。强有力的疫苗接种计划以及快速和准确的实验室诊断在有效控制和根除该疾病方面发挥着重要作用。此外，对正在传播的猪繁殖与呼吸综合征病毒的基因分析有助于确定该病毒在邻国之间传播的流行病学联系。在这里，我们开发了一种改进的聚合酶链式反应(PCR)，它可以检测到低病毒载量，并有助于遗传鉴定，特别是在资源有限的环境中。小反刍动物疫病(PPR)是由PPR病毒(PPRV)引起的一种高度传染性的小型反刍动物病毒性疾病。PPR在亚洲、中东和非洲大片地区流行，目前的目标是到2030年在全球根除PPR。该病毒以四种不同的谱系存在，通常局限于特定的地理区域。然而，最近关于猪繁殖与呼吸综合征病毒传播的报告，特别是向无感染国家和以前的猪繁殖与呼吸综合征流行地区传播的IV型病毒，值得注意。快速、准确的实验室诊断和关于其与病毒传播的流行病学联系的报告在有效控制和根除该疾病方面发挥着重要作用。目前，PPR的诊断主要采用分子生物学方法，包括常规逆转录聚合酶链式反应(RT-PCR)和实时定量RT-PCR(RT-qPCR)，核衣壳基因部分序列测定用于病毒谱系鉴定。然而，如果动物在感染初期排泄的病毒水平很低，或者如果PPRV在长途运输样本到参考实验室的过程中被降解，那么很难对遗传物质进行诊断和测序。本研究建立了一种新的针对N蛋白基因的套式RT-PCR检测猪繁殖与呼吸综合征病毒核酸的方法，并与现有的常规RT-PCR方法进行了比较，同时还提供了高质量的DNA，可用于测序以鉴定循环谱系。使用繁殖的猪繁殖病毒的细胞培养、临床感染动物的现场样本和涵盖所有四种猪繁殖与呼吸综合征病毒(I-IV)谱系的实验感染动物的样本对该检测进行了评估。该分析提供了一种简单、准确、快速和经济高效的PPR诊断和部分基因组测序的解决方案，可用于资源有限的环境。
Peste des petits ruminants (PPR) is a highly contagious and economically important viral disease of small ruminants and a large number of wildlife species. The causative agent, PPR virus (PPRV) is mainly circulating across large areas in Africa, the Middle East and Asia and is, fortunately, currently subject to a global eradication program. A robust vaccination program along with a rapid and accurate laboratory diagnosis play a major role in the effective control and eradication of the disease. Furthermore, a genetic analysis of the circulating PPRV can help to identify the epidemiological linkage of the virus spread between neighboring countries. Here, we have developed an improved polymerase chain reaction (PCR) that can detect low viral loads and help in genetic identification, particularly in resource-limited settings. Peste des petits ruminants (PPR) is a highly contagious viral disease of small ruminants caused by PPR virus (PPRV). PPR is endemic in Asia, the Middle East and across large areas of Africa and is currently targeted for global eradication by 2030. The virus exists as four different lineages that are usually limited to specific geographical areas. However, recent reports of spread of PPRV, in particular of lineage IV viruses to infection-free countries and previously PPR endemic areas are noteworthy. A rapid and accurate laboratory diagnosis and reports on its epidemiological linkage for virus spread play a major role in the effective control and eradication of the disease. Currently, molecular assays, including conventional reverse transcription-polymerase chain reaction (RT-PCR) and real-time RT-PCR (RT-qPCR) are usually used for diagnosis of PPR while the sequencing of part of the nucleocapsid gene is usually carried out for the viral lineage identification. However, it is difficult to diagnose and sequence the genetic material if the animal excreted a low level of virus at the initial stage of infection or if the PPRV is degraded during the long-distance transportation of samples to the reference laboratories. This study describes the development of a novel nested RT-PCR assay for the detection of the PPRV nucleic acid by targeting the N-protein gene, compares the performance of the assay with the existing conventional RT-PCR and also provides good-quality DNA suitable for sequencing in order to identify circulating lineages. The assay was evaluated using cell culture propagated PPRVs, field samples from clinically infected animals and samples from experimentally infected animals encompassing all four lineages (I–IV) of PPRV. This assay provides a solution with an easy, accurate, rapid and cost-effective PPR diagnostic and partial genome sequencing for use in resource-limited settings.
登录
查看更多内容
影响因子：
4.3
作者：
Clarke BD;Islam MR;Yusuf MA;Mahapatra M;Parida S
通讯作者：
Parida S
DOI：
10.3390/v13112144
发表时间：
2021-10-25
期刊：
Viruses
影响因子：
--
作者：
Mahapatra M;Pachauri R;Subramaniam S;Banyard AC;ChandraSekar S;Ramakrishnan MA;Njeumi F;Muthuchelvan D;Parida S
通讯作者：
Parida S
DOI：
10.3390/ani11102976
发表时间：
2021-10-15
期刊：
Animals : an open access journal from MDPI
影响因子：
--
作者：
Kinimi E;Mahapatra M;Kgotlele T;Makange MR;Tennakoon C;Njeumi F;Odongo S;Muyldermans S;Kock R;Parida S;Rweyemamu M;Misinzo G
通讯作者：
Misinzo G
影响因子：
4.7
作者：
Mahapatra, Mana;Howson, Emma;Parida, Satya
通讯作者：
Parida, Satya
影响因子：
4.3
作者：
Niyokwishimira, Alfred;Baziki, Jean de D.;Bodjo, Sanne Charles
通讯作者：
Bodjo, Sanne Charles