The globally invasive cassava mealybug Phenacoccus manihoti Matile-Ferrero is a pernicious pest of cassava,and its recent introduction into Asia has raised considerable alarm.To slow or prevent further invasion,an acc...The globally invasive cassava mealybug Phenacoccus manihoti Matile-Ferrero is a pernicious pest of cassava,and its recent introduction into Asia has raised considerable alarm.To slow or prevent further invasion,an accurate,simple,and developmental-stage-independent detection method for P.manihoti is required.In the present study,a PCR method based on a species-specific mitochondrial DNA cytochrome oxidase I(SS-COI)marker was developed for rapid identification of P.manihoti.One pair of SS-COI primers(PMSSZW-1F and PMSSZW-1R)was designed based on sequence variations in the COI gene among P.manihoti and related mealybug species.Specificity of the primer pair was validated on 21 closely related species.Sensitivity tests were performed on four immature developmental stages and female adults.Efficacy tests demonstrated that at the relatively low concentration of(135.2±14.7)pgresuspended DNA,the specific fragment was detected in all replicates.Furthermore,the SS-COI primer pair was assayed on three populations of P.manihoti from major exporting countries of cassava.The PCR assay was proved to be a rapid,simple,and reliable molecular measure for the identification of P.manihoti.This tool will be useful for quarantine,monitoring,and management of this invasive pest.展开更多
Cassava bacterial blight (CBB) caused by Xanthomonas axonopodis pv. manihotis has been reported in several African countries since 1970. Knowledge of the virulence and diversity of Xanthomonas axonopodis pv. manihotis...Cassava bacterial blight (CBB) caused by Xanthomonas axonopodis pv. manihotis has been reported in several African countries since 1970. Knowledge of the virulence and diversity of Xanthomonas axonopodis pv. manihotis strains is important for an integrated control of CBB. The main objective of the present study was to characterize strains of Xanthomonas axonopodis collected from various regions in the DR-Congo. There was variability among strains for shape (form), contour (margin) and elevation. Bacterial cell size for the strains analyzed varied from 24.1 μm × 11.3 μm to 11.4 μm × 4.2 μm. All the Xanthomonas axonopodis pv. manihotis strains but one was motile. Two distinctive groups were identified based on radial growth of their colonies. The first group grows faster (7.8-10.5 mm/d) compared to the second group (4.8-6.9 mm/d). Five strains (Gandajika, Inera/Stat, Kansasa, Mulumba and Musakatshi) were classified as virulent with a damage rating ≤ 1 and four were aggressive (Luputa, M'vuazi, Boketa and Kiyaka) with a damage rating > 1. Significant differences were also observed among strains for disease onset, incidence and plant mortality. The highest incidence (33%) of bacterial blight 21 days after infestation (DAI) resulted from the Boketa strain inoculation and the lowest (0 % disease incidence) from INERA/STAT and Musakatshi strains. There was no clear association between geographic origin of the strains and their aggressiveness.展开更多
建立了一种木薯细菌性萎蔫病菌的环介导恒温扩增快速检测方法,为木薯细菌性萎蔫病的快速检测提供有力的技术支持。针对木薯细菌性萎蔫病菌TAL效应器蛋白质(pthBXam)靶序列的6个位点设计4条特异性引物,并对反应温度和内引物浓度等参数进...建立了一种木薯细菌性萎蔫病菌的环介导恒温扩增快速检测方法,为木薯细菌性萎蔫病的快速检测提供有力的技术支持。针对木薯细菌性萎蔫病菌TAL效应器蛋白质(pthBXam)靶序列的6个位点设计4条特异性引物,并对反应温度和内引物浓度等参数进行了优化,设计的引物与试验中提供的其他黄单胞近缘种都没有扩增反应,表现了较好的特异性。LAMP方法对木薯细菌性萎蔫病菌菌株DNA的检测下限为1pg/μL,比常规PCR灵敏度高100倍。该方法采用SYBR Green I染料法对扩增产物闭管检测,裸眼观察颜色变化判断反应结果,能快速、准确地对田间样品进行检测,没有出现假阳性和假阴性。与其他检测方法相比,LAMP方法检测时间短,效率高,降低了设备投入,易于操作,适合木薯细菌性萎蔫病菌的现场检疫和大规模监测。展开更多
基金supported by the National Key R&D Program of China(2017YFC1200600,2016YFC1201200 and 2015BAD08A16)the Science and Technology Innovation Program of CAAS(caascx-2013-2018-IAS)
文摘The globally invasive cassava mealybug Phenacoccus manihoti Matile-Ferrero is a pernicious pest of cassava,and its recent introduction into Asia has raised considerable alarm.To slow or prevent further invasion,an accurate,simple,and developmental-stage-independent detection method for P.manihoti is required.In the present study,a PCR method based on a species-specific mitochondrial DNA cytochrome oxidase I(SS-COI)marker was developed for rapid identification of P.manihoti.One pair of SS-COI primers(PMSSZW-1F and PMSSZW-1R)was designed based on sequence variations in the COI gene among P.manihoti and related mealybug species.Specificity of the primer pair was validated on 21 closely related species.Sensitivity tests were performed on four immature developmental stages and female adults.Efficacy tests demonstrated that at the relatively low concentration of(135.2±14.7)pgresuspended DNA,the specific fragment was detected in all replicates.Furthermore,the SS-COI primer pair was assayed on three populations of P.manihoti from major exporting countries of cassava.The PCR assay was proved to be a rapid,simple,and reliable molecular measure for the identification of P.manihoti.This tool will be useful for quarantine,monitoring,and management of this invasive pest.
文摘Cassava bacterial blight (CBB) caused by Xanthomonas axonopodis pv. manihotis has been reported in several African countries since 1970. Knowledge of the virulence and diversity of Xanthomonas axonopodis pv. manihotis strains is important for an integrated control of CBB. The main objective of the present study was to characterize strains of Xanthomonas axonopodis collected from various regions in the DR-Congo. There was variability among strains for shape (form), contour (margin) and elevation. Bacterial cell size for the strains analyzed varied from 24.1 μm × 11.3 μm to 11.4 μm × 4.2 μm. All the Xanthomonas axonopodis pv. manihotis strains but one was motile. Two distinctive groups were identified based on radial growth of their colonies. The first group grows faster (7.8-10.5 mm/d) compared to the second group (4.8-6.9 mm/d). Five strains (Gandajika, Inera/Stat, Kansasa, Mulumba and Musakatshi) were classified as virulent with a damage rating ≤ 1 and four were aggressive (Luputa, M'vuazi, Boketa and Kiyaka) with a damage rating > 1. Significant differences were also observed among strains for disease onset, incidence and plant mortality. The highest incidence (33%) of bacterial blight 21 days after infestation (DAI) resulted from the Boketa strain inoculation and the lowest (0 % disease incidence) from INERA/STAT and Musakatshi strains. There was no clear association between geographic origin of the strains and their aggressiveness.
文摘建立了一种木薯细菌性萎蔫病菌的环介导恒温扩增快速检测方法,为木薯细菌性萎蔫病的快速检测提供有力的技术支持。针对木薯细菌性萎蔫病菌TAL效应器蛋白质(pthBXam)靶序列的6个位点设计4条特异性引物,并对反应温度和内引物浓度等参数进行了优化,设计的引物与试验中提供的其他黄单胞近缘种都没有扩增反应,表现了较好的特异性。LAMP方法对木薯细菌性萎蔫病菌菌株DNA的检测下限为1pg/μL,比常规PCR灵敏度高100倍。该方法采用SYBR Green I染料法对扩增产物闭管检测,裸眼观察颜色变化判断反应结果,能快速、准确地对田间样品进行检测,没有出现假阳性和假阴性。与其他检测方法相比,LAMP方法检测时间短,效率高,降低了设备投入,易于操作,适合木薯细菌性萎蔫病菌的现场检疫和大规模监测。
