Swine influenza A virus（swine IAV） circulates worldwide in pigs and poses a serious public health threat, as evidenced by the 2009 H1N1 influenza pandemic. Among multiple subtypes/lineages of swine influenza A virus...Swine influenza A virus（swine IAV） circulates worldwide in pigs and poses a serious public health threat, as evidenced by the 2009 H1N1 influenza pandemic. Among multiple subtypes/lineages of swine influenza A viruses, European avian-like（EA） H1N1 swine IAV has been dominant since 2005 in China and caused infections in humans in 2010. Highly sensitive and specific methods of detection are required to differentiate EA H1N1 swine IAVs from viruses belonging to other lineages and subtypes. In this study, a nested reverse transcription（RT）-PCR assay was developed to detect EA H1 swine IAVs. Two primer sets（outer and inner） were designed specifically to target the viral hemagglutinin genes. Specific PCR products were obtained from all tested EA H1N1 swine IAV isolates, but not from other lineages of H1 swine IAVs, other subtypes of swine IAVs, or other infectious swine viruses. The sensitivity of the nested RT-PCR was improved to 1 plaque forming unit（PFU） m L^（-1） which was over 10~4 PFU m L^（-1） for a previously established multiplex RT-PCR method. The nested RT-PCR results obtained from screening 365 clinical samples were consistent with those obtained using conventional virus isolation methods combined with sequencing. Thus, the nested RT-PCR assay reported herein is more sensitive and suitable for the diagnosis of clinical infections and surveillance of EA H1 swine IAVs in pigs and humans.展开更多
OBJECTIVES: To detect blood AFPmRNA in the nude mice bearing with human hepatocellular carcinoma (HCC) by using nested reverse transcriptase polymerase chain reaction (nested RT-PCR), and assess its significance in HC...OBJECTIVES: To detect blood AFPmRNA in the nude mice bearing with human hepatocellular carcinoma (HCC) by using nested reverse transcriptase polymerase chain reaction (nested RT-PCR), and assess its significance in HCC distant metastasis. METHOD: We detected 20 blood samples from the nude mice bearing with human HCC by nested RT-PCR to find out AFPmRNA. RESULT: AFPmRNA was detected in 6 blood samples from the nude mice hearing with human HCC (30. 0%), in which 4 mice developed distant metastasis. CONCLUSION: AFPmRNA may be used as an efficient and sensitive marker to detect blood spread of HCC cells. It can predict the occurrence of HCC distant metastasis.展开更多
Reverse transcription polymerase chain reaction (RT-PCR) was used for the detection of classical swine fever virus (CSFV) in blood and tissue samples of field cases and experimentally inoculated pigs. The distribution...Reverse transcription polymerase chain reaction (RT-PCR) was used for the detection of classical swine fever virus (CSFV) in blood and tissue samples of field cases and experimentally inoculated pigs. The distribution of CSFV in different organ samples showed some discrepancies in infected pigs. Four weaner pigs were inoculated with C-strain vaccine virus, then samples of spleen, tonsil, lung, mesenteric lymph node, kidney and brain were collected after slaughter and tested for E2 and NS5B genes using one-step RT-PCR and nested RT-PCR. Using the same method, 12 field cases were simultaneously studied. A discrepancy of CSFV in different samples was found upon detecting the target gene. The most reliable diagnostic organs were spleen and tonsil, and the nested RT-PCR assay provided a highly sensitive and specific method with comparable performance to the one-step RT-PCR assay.展开更多
背景A群轮状病毒(group A rotavirus,RVA)是导致婴幼儿及幼龄动物发生腹泻的主要人兽共患病病原之一,具有较强的传染性。目的建立一种快速、特异检测猪、牛、羊、人等物种RVA的实时荧光定量RT-PCR检测方法。方法基于GenBank中的猪、牛...背景A群轮状病毒(group A rotavirus,RVA)是导致婴幼儿及幼龄动物发生腹泻的主要人兽共患病病原之一,具有较强的传染性。目的建立一种快速、特异检测猪、牛、羊、人等物种RVA的实时荧光定量RT-PCR检测方法。方法基于GenBank中的猪、牛、羊和人源RVA NSP5基因保守序列,设计1对特异性引物、1条探针,该探针带有TaqMan-MGB发光基团,优化最适退火温度(Tm)、最适引物和探针浓度,建立一种可以检测猪、牛、羊和人等物种RVA的实时荧光定量RT-PCR检测方法。结果建立的RVA荧光定量RT-PCR方法最佳退火温度为51℃;最适引物和探针浓度均为0.4μmol/L;敏感性高,最低检测限可达3.24 copies/μL;特异性强,与猪、牛、羊等常见病原无检测交叉反应;重复性好,其批内和批间试验变异系数分别为0.063%-0.812%和0.612%-1.016%,可以快速、特异检测猪、牛、羊样品和人轮状病毒,有很好的临床适用性。结论本实验建立了一种检测多物种RVA的实时荧光定量RT-PCR方法,为猪、牛、羊和人等多物种的RVA流行病学调查和早期诊断提供了良好的技术支撑。展开更多
为了实现对存在传入风险的南非2型(South African type 2, SAT2)口蹄疫病毒的早期发现、精准鉴定和有效预警,本研究基于SAT2型口蹄疫病毒毒株IRAN/2024和ETH/2022完整VP1基因序列,构建了pUC57-IRAN-VP1、pUC57-ETH-VP1质粒;参考SAT2型...为了实现对存在传入风险的南非2型(South African type 2, SAT2)口蹄疫病毒的早期发现、精准鉴定和有效预警,本研究基于SAT2型口蹄疫病毒毒株IRAN/2024和ETH/2022完整VP1基因序列,构建了pUC57-IRAN-VP1、pUC57-ETH-VP1质粒;参考SAT2型口蹄疫病毒VP1基因序列设计并筛选特异性引物,以所构建的质粒为模板,建立了SAT2型口蹄疫病毒特异性RT-PCR检测方法,并开展敏感性试验、特异性试验。敏感性试验结果显示,该方法可以检测质量浓度低至1 pg/mL的质粒DNA。特异性试验结果显示,该方法对伪狂犬病毒、猪繁殖与呼吸综合征病毒、乙型脑炎病毒、猪瘟病毒、1型蓝舌病病毒、牛病毒性腹泻病毒、山羊痘病毒、阿卡斑病毒、流行性出血热病毒等常见病毒的核酸,以及参试的O型和A型口蹄疫病毒(PanAsia、Cathay、Mya98、Ind2001-1、Ind2001-2、AKT-Ⅲ、Sea-97毒株)核酸均无交叉反应。应用该方法对2023年云南边境地区50份牛食道-咽部分泌物样品进行核酸检测,检测结果与RT-qPCR检测结果一致。本研究建立的SAT2型口蹄疫病毒特异性RT-PCR检测方法具有一定的实用性,为口蹄疫疫情防控提供技术支撑。展开更多
Sparse array design has significant implications for improving the accuracy of direction of arrival(DOA)estimation of non-circular(NC)signals.We propose an extended nested array with a filled sensor(ENAFS)based on the...Sparse array design has significant implications for improving the accuracy of direction of arrival(DOA)estimation of non-circular(NC)signals.We propose an extended nested array with a filled sensor(ENAFS)based on the hole-filling strategy.Specifically,we first introduce the improved nested array(INA)and prove its properties.Subsequently,we extend the sum-difference coarray(SDCA)by adding an additional sensor to fill the holes.Thus the larger uniform degrees of freedom(uDOFs)and virtual array aperture(VAA)can be abtained,and the ENAFS is designed.Finally,the simulation results are given to verify the superiority of the proposed ENAFS in terms of DOF,mutual coupling and estimation performance.展开更多
Nest predation is a critical driver of avian reproductive success,yet its response to urbanization remains controversial.Using infrared camera traps to monitor 133 artificial nests and 32 natural nests of the Spotted ...Nest predation is a critical driver of avian reproductive success,yet its response to urbanization remains controversial.Using infrared camera traps to monitor 133 artificial nests and 32 natural nests of the Spotted Dove(Spilopelia chinensis) along an urbanization gradient in Nanchang and Shangrao,China,from 2021 to 2023,we examined spatiotemporal variation in predation pressure.We deconstructed predation pressure into risk(nest fate and survival time) and pattern(predation frequency,predator size,and predator-nest interaction duration).We found that higher urbanization and better nest concealment significantly reduced predation risk.The composition of the predator community did not change along the urbanization gradient,suggesting that the reduced nest predation risk in cities is likely driven by behavioral factors,such as the availability of alternative food or human shield effect,rather than by the filtering of predator species.In contrast,predation patterns were primarily moderated by season.The off-peak breeding season was char acterized by more frequent but less severe predation events involving smaller predators and had shorter predator-nest interaction durations compared to the peak season.This seasonal shift opens a breeding window during the off-peak season,when lethal nest predation risk is reduced.The Spotted Dove's unique life-history traits enable it to capitalize on this opportunity by balancing environmental costs,enhancing its reproductive success and facilitating its colonization of urban environments.展开更多
本研究以日本黄瓜绿斑驳花叶病毒(kyuri green mottle mosaic virus,KGMMV)为材料,设计1对常规RT-PCR检测引物KGCPN-F/KGCPN-R和1组引物/探针KGM-F/KGM-R/KGM-P,分析测试了2对已发表KGMMV常规RT-PCR检测引物KGCP-F/KGCP-R和KGMP-F/KGMP...本研究以日本黄瓜绿斑驳花叶病毒(kyuri green mottle mosaic virus,KGMMV)为材料,设计1对常规RT-PCR检测引物KGCPN-F/KGCPN-R和1组引物/探针KGM-F/KGM-R/KGM-P,分析测试了2对已发表KGMMV常规RT-PCR检测引物KGCP-F/KGCP-R和KGMP-F/KGMP-R,建立了KGMMV的常规和实时荧光RT-PCR检测方法。结果表明,引物KGCP-F/KGCP-R扩增KGMMV时出现预期大小的条带,扩增黄瓜绿斑驳花叶病毒(cucumber green mottle mosaic virus,CGMMV)时出现非常微弱的条带;引物KGCPN-F/KGCPN-R扩增KGMMV时出现预期大小的条带,扩增小西葫芦绿斑驳花叶病毒(zucchini green mottle mosaic virus,ZGMMV)时出现比预期稍大的条带,通过对PCR产物进行序列测定和分析比对可准确鉴定KGMMV。KGCP-F/KGCP-R和KGCPN-F/KGCPN-R的相对灵敏度分别为10^(-6)和10^(-5)稀释度,适用于KGMMV的常规RT-PCR检测。基于引物探针KGM-F/KGM-R/KGM-P建立的KGMMV实时荧光RT-PCR检测方法能特异性检出KGMMV,相对灵敏度达10^(-7)稀释度,分别比2对常规RT-PCR检测引物高10倍和100倍,适用于瓜类种子中KGMMV的快速检测。展开更多
基金supported by the National High-Tech R&D Program of China (2012AA101303)
文摘Swine influenza A virus（swine IAV） circulates worldwide in pigs and poses a serious public health threat, as evidenced by the 2009 H1N1 influenza pandemic. Among multiple subtypes/lineages of swine influenza A viruses, European avian-like（EA） H1N1 swine IAV has been dominant since 2005 in China and caused infections in humans in 2010. Highly sensitive and specific methods of detection are required to differentiate EA H1N1 swine IAVs from viruses belonging to other lineages and subtypes. In this study, a nested reverse transcription（RT）-PCR assay was developed to detect EA H1 swine IAVs. Two primer sets（outer and inner） were designed specifically to target the viral hemagglutinin genes. Specific PCR products were obtained from all tested EA H1N1 swine IAV isolates, but not from other lineages of H1 swine IAVs, other subtypes of swine IAVs, or other infectious swine viruses. The sensitivity of the nested RT-PCR was improved to 1 plaque forming unit（PFU） m L^（-1） which was over 10~4 PFU m L^（-1） for a previously established multiplex RT-PCR method. The nested RT-PCR results obtained from screening 365 clinical samples were consistent with those obtained using conventional virus isolation methods combined with sequencing. Thus, the nested RT-PCR assay reported herein is more sensitive and suitable for the diagnosis of clinical infections and surveillance of EA H1 swine IAVs in pigs and humans.
文摘OBJECTIVES: To detect blood AFPmRNA in the nude mice bearing with human hepatocellular carcinoma (HCC) by using nested reverse transcriptase polymerase chain reaction (nested RT-PCR), and assess its significance in HCC distant metastasis. METHOD: We detected 20 blood samples from the nude mice bearing with human HCC by nested RT-PCR to find out AFPmRNA. RESULT: AFPmRNA was detected in 6 blood samples from the nude mice hearing with human HCC (30. 0%), in which 4 mice developed distant metastasis. CONCLUSION: AFPmRNA may be used as an efficient and sensitive marker to detect blood spread of HCC cells. It can predict the occurrence of HCC distant metastasis.
基金supported by the grants from the National"863" Programme (2006AA10A2041)Development Programme of Science and Technology,Chengguan District of Gansu province (08-5-4)
文摘Reverse transcription polymerase chain reaction (RT-PCR) was used for the detection of classical swine fever virus (CSFV) in blood and tissue samples of field cases and experimentally inoculated pigs. The distribution of CSFV in different organ samples showed some discrepancies in infected pigs. Four weaner pigs were inoculated with C-strain vaccine virus, then samples of spleen, tonsil, lung, mesenteric lymph node, kidney and brain were collected after slaughter and tested for E2 and NS5B genes using one-step RT-PCR and nested RT-PCR. Using the same method, 12 field cases were simultaneously studied. A discrepancy of CSFV in different samples was found upon detecting the target gene. The most reliable diagnostic organs were spleen and tonsil, and the nested RT-PCR assay provided a highly sensitive and specific method with comparable performance to the one-step RT-PCR assay.
文摘背景A群轮状病毒(group A rotavirus,RVA)是导致婴幼儿及幼龄动物发生腹泻的主要人兽共患病病原之一,具有较强的传染性。目的建立一种快速、特异检测猪、牛、羊、人等物种RVA的实时荧光定量RT-PCR检测方法。方法基于GenBank中的猪、牛、羊和人源RVA NSP5基因保守序列,设计1对特异性引物、1条探针,该探针带有TaqMan-MGB发光基团,优化最适退火温度(Tm)、最适引物和探针浓度,建立一种可以检测猪、牛、羊和人等物种RVA的实时荧光定量RT-PCR检测方法。结果建立的RVA荧光定量RT-PCR方法最佳退火温度为51℃;最适引物和探针浓度均为0.4μmol/L;敏感性高,最低检测限可达3.24 copies/μL;特异性强,与猪、牛、羊等常见病原无检测交叉反应;重复性好,其批内和批间试验变异系数分别为0.063%-0.812%和0.612%-1.016%,可以快速、特异检测猪、牛、羊样品和人轮状病毒,有很好的临床适用性。结论本实验建立了一种检测多物种RVA的实时荧光定量RT-PCR方法,为猪、牛、羊和人等多物种的RVA流行病学调查和早期诊断提供了良好的技术支撑。
基金supported by China National Science Foundations(Nos.62371225,62371227)。
文摘Sparse array design has significant implications for improving the accuracy of direction of arrival(DOA)estimation of non-circular(NC)signals.We propose an extended nested array with a filled sensor(ENAFS)based on the hole-filling strategy.Specifically,we first introduce the improved nested array(INA)and prove its properties.Subsequently,we extend the sum-difference coarray(SDCA)by adding an additional sensor to fill the holes.Thus the larger uniform degrees of freedom(uDOFs)and virtual array aperture(VAA)can be abtained,and the ENAFS is designed.Finally,the simulation results are given to verify the superiority of the proposed ENAFS in terms of DOF,mutual coupling and estimation performance.
基金financially supported by the Jiangxi Provincial Natural Science Foundation,China (No. 20224ACB205006)。
文摘Nest predation is a critical driver of avian reproductive success,yet its response to urbanization remains controversial.Using infrared camera traps to monitor 133 artificial nests and 32 natural nests of the Spotted Dove(Spilopelia chinensis) along an urbanization gradient in Nanchang and Shangrao,China,from 2021 to 2023,we examined spatiotemporal variation in predation pressure.We deconstructed predation pressure into risk(nest fate and survival time) and pattern(predation frequency,predator size,and predator-nest interaction duration).We found that higher urbanization and better nest concealment significantly reduced predation risk.The composition of the predator community did not change along the urbanization gradient,suggesting that the reduced nest predation risk in cities is likely driven by behavioral factors,such as the availability of alternative food or human shield effect,rather than by the filtering of predator species.In contrast,predation patterns were primarily moderated by season.The off-peak breeding season was char acterized by more frequent but less severe predation events involving smaller predators and had shorter predator-nest interaction durations compared to the peak season.This seasonal shift opens a breeding window during the off-peak season,when lethal nest predation risk is reduced.The Spotted Dove's unique life-history traits enable it to capitalize on this opportunity by balancing environmental costs,enhancing its reproductive success and facilitating its colonization of urban environments.
文摘本研究以日本黄瓜绿斑驳花叶病毒(kyuri green mottle mosaic virus,KGMMV)为材料,设计1对常规RT-PCR检测引物KGCPN-F/KGCPN-R和1组引物/探针KGM-F/KGM-R/KGM-P,分析测试了2对已发表KGMMV常规RT-PCR检测引物KGCP-F/KGCP-R和KGMP-F/KGMP-R,建立了KGMMV的常规和实时荧光RT-PCR检测方法。结果表明,引物KGCP-F/KGCP-R扩增KGMMV时出现预期大小的条带,扩增黄瓜绿斑驳花叶病毒(cucumber green mottle mosaic virus,CGMMV)时出现非常微弱的条带;引物KGCPN-F/KGCPN-R扩增KGMMV时出现预期大小的条带,扩增小西葫芦绿斑驳花叶病毒(zucchini green mottle mosaic virus,ZGMMV)时出现比预期稍大的条带,通过对PCR产物进行序列测定和分析比对可准确鉴定KGMMV。KGCP-F/KGCP-R和KGCPN-F/KGCPN-R的相对灵敏度分别为10^(-6)和10^(-5)稀释度,适用于KGMMV的常规RT-PCR检测。基于引物探针KGM-F/KGM-R/KGM-P建立的KGMMV实时荧光RT-PCR检测方法能特异性检出KGMMV,相对灵敏度达10^(-7)稀释度,分别比2对常规RT-PCR检测引物高10倍和100倍,适用于瓜类种子中KGMMV的快速检测。
