Rapid detection and identification of the bacterium Pantoea stewartii in maize by TaqMan real-time PCR assay targeting the cpsD gene

Aims: The development and evaluation of a sensitive and specific TaqMan^® real-time polymerase chain reaction (PCR) for the detection and identification of Pantoea stewartii on maize. Methods and Results: A TaqMan^®-based real-time PCR assay targeting the cpsD gene enabling specific detection of P. stewartii in maize leaves and seeds was developed. Under optimal conditions, the selected primers and probe were specific for the detection of all 14 reference P. stewartii strains by real-time PCR. The 32 non-Panteoa and eight other Pantoea strains tested negative. The TaqMan^® PCR assay detected 1 pg of purified DNA and 10⁴P. stewartii colony forming units per millilitre (10 cells per reaction) in pure cultures consisting of 92·0% intact (viable) cells. Direct processing of leaf lesions and seeds by the real-time PCR detected 10 and 50 P. stewartii cells per reaction respectively. TaqMan^® real-time PCR results were validated by dilution plating of macerates and PCR-based subcloning followed by DNA sequencing. Conclusions: The real-time PCR assay described is a rapid, reliable and more sensitive tool for the detection of P. stewartii. Significance and Impact of the study: This real-time PCR assay would avoid false-negative results and reduce the time required for certifying maize seed shipments.     

玉米细菌性枯萎病及其病原菌的检测技术

玉米细菌性枯萎病是影响玉米生产的重要病害 ,该病害通过带菌的种子进行远距离传播 ,对该病菌的检测成为防止和控制该病害的重要手段。介绍现有的检测该病菌的各种方法 ,即黑色素选择培养法、double sandwichELISA以及RAPD PCR ,LCR PCR ,Nested PCR ,multiplePCR和荧光实时PCR等分子生物学检测方法 ,并对这些方法的特性进行比较和探讨。     

玉米细菌性枯萎病菌改良Dot-ELISA检测研究

通过激光切割技术加工出硝酸纤维素膜小圆片并粘贴在已打孔的塑料胶条上,制成包含8个圆片的NCM条(NCM test strip),进一步在NCM条上建立了玉米细菌性枯萎病菌的Dot-ELISA检测方法.研究发现,在NCM圆片上的Dot-ELISA检测灵敏度与点样量密切相关,采用10 μL/点的加样量比通常的1 μL/点可提高检测灵敏度10-100倍;间接Dot-ELISA的检测灵敏度是双抗夹心dot-ELISA的10倍,该结果进一步在微孔板ELISA的检测中得到证实.玉米细菌性枯萎病菌的改良Dot-ELISA检测是一种较为灵敏、快速、稳定、规范的实验方法,为进一步研究该病菌的微流控芯片斑点免疫检测方法奠定了前期工作基础.     

Hypothesis tests for and against a simple order among proportions estimated by pooled testing

The use of pooled testing as a means of estimating the prevalence of rare traits has received considerable attention in recent years, particularly in the areas of public health, genetics, animal-disease assessment, and plant pathology. In pooled-testing applications, observations are made on pools of individuals amalgamated together. In this paper, we examine order-restricted hypothesis tests involving k >2 binomial proportions estimated by pooled testing, extending the earlier work of Tebbs and Swallow (2003, Biometrika, 90, 471-477 and Biometrical Journal, 45, 618-630). In particular, we focus on (i) testing the equality of proportions versus an isotonic alternative and (ii) testing for a violation of isotonicity. We propose new tests for each scenario and provide results which characterize the small-sample performance of our procedures. We illustrate our methods using two data sets; one from an observational HIV study and one from an agricultural experiment.     

温度对尖裸鲤胚胎发育及其仔稚鱼生长性状的影响

采用实验生态学方法研究温度对尖裸鲤（Oxygymnocypris stewartii）胚胎发育及仔稚鱼生长性状的影响，旨在掌握温度对其早期发育的影响。结果表明，随着温度的升高，胚胎的孵化时间缩短，发育速度加快。在平均水温5 ℃、8 ℃、11 ℃、14 ℃和17 ℃下，尖裸鲤的胚胎孵化时间分别为530.78 h、366.12 h、214.22 h、220.63 h、153.95 h，温度和孵化时间呈幂函数关系，有效积温在水温为11 ℃时最低，为2 356.4 h ·℃。尖裸鲤胚胎不同发育时期在不同温度下的累计时间均呈现指数函数关系。随着温度的升高，孵化率呈现先降低后升高的趋势，水温范围在11 ~ 17 ℃时，温度系数Q10值和Q13值最接近2。尖裸鲤初孵仔鱼全长与温度间呈现三次多项式函数，全长与温度之间不存在显著性关系；而初孵仔鱼卵黄囊体积与温度间呈现显著性的二次多项式函数。综合各项指标表明，尖裸鲤胚胎的适宜孵化水温为11 ~ 17 ℃，仔稚鱼的适宜生长水温为14 ~ 17 ℃。