Molecular prevalence,risk factors assessment and haemato-biochemical alterations in hepatozoonosis in dogs from Punjab,India

Hepatozoonosis caused by Hepatozoon canis is an important tick-borne disease of dogs in tropical and sub-tropical regions throughout the world. In the present study evaluation of blood samples collected from 225 dogs presented at Small Animal Clinics, GADVASU, Ludhiana, Punjab (India) was done for the presence of H. canis by PCR based assay targeting a portion of 18S rRNA gene. Of the total samples subjected to PCR, an amplicon of 666 bp was detected in 13.78% samples whereas, routine blood smear examination revealed gamonts in 5.78% samples. Furthermore, prevalence of H. canis infection was found to be significantly associated with season, being highest in summer and lowest in winter while other risk factors e.g. age, sex and breed showed non-significant association. In terms of various clinico-pathological parameters, significant drop in haemoglobin, total red blood cell count, packed cell volume and lymphocytes were recorded in positive cases whereas the total white blood cell count was non-significantly increased. The haematological alterations in the positive cases were lymphopenia, anaemia, thrombocytopenia, relative neutrophilia, neutrophilic leucocytosis, eosinophilia, monocytosis and lymphocytosis while the biochemical profile revealed hypoproteinemia and increased levels of blood urea nitrogen and creatinine (in positive cases) pointing towards renal failure.     

利用QuantStudioTM 3D数字PCR分析转基因玉米MON863含量

QuantStudioTM 3D数字PCR (QuantStudioTM 3D digital PCR,3D-dPCR)是一种基于超高密度亲疏水微孔芯片实现数字PCR分液原理的新型核酸绝对定量平台,在转基因生物定量领域具有极大的应用前景.本研究基于3D-dPCR平台,以转基因玉米(Zea mays)MON863混合样品为例,建立基于单重和双重数字PCR体系的转基因生物(genetically modified organisms,GMOs)含量分析方法.与传统qRT-PCR比较发现,在缺乏样品纯度、纯合度信息的情况下,数字PCR能够较好地排除这些因素的影响,测定准确的量值.研究结果表明,QuantStudioTM 3D数字PCR是一种适用于转基因生物含量分析的精确定量方法,还可反映转基因玉米种子的基因型.本研究基于3D-dPCR建立的转基因玉米MON863单重和双重定量方法为转基因检测提供了新的方法和参考.     

Prevalence,molecular characterization and risk factor analysis of Ehrlichia canis and Anaplasma platys in domestic dogs from Paraguay

This is the first study to investigate the prevalence and risk factors associated with Ehrlichia canis and Anaplasma platys positivity in dogs from Paraguay. Conventional PCR assays for the E. canis 16SrRNA gene and A. platys p44 gene were carried out in blood samples from 384 dogs from Asunción city, Paraguay. Sequencing and phylogenetic analysis were performed in selected positive E. canis and (16SrRNA gene) and A. platys (16S and p44 genes) samples. The overall prevalence of E. canis and A. platys in dogs in Paraguay was 10.41% (40/384) and 10.67% (41/384), respectively. Older dogs without veterinary care had higher odds for E. canis positivity and a higher number of dogs in the same household, as well as absence of anti-tick treatment were considered risk factors for A. platys. Ehrlichia canis and A. platys circulate in the dog population from Asunción, and are described for the first time in Paraguay.     

水稻复合农艺性状QTL剖析

为了解水稻复合性状的数量性状基因座(QTL),在利用单片段代换系进行QTL鉴定的基础上,剖析了水稻株高QTL与主茎高和穗长QTL,主茎高与倒一节间长、倒二节间长、倒三节间长和倒四及以下节间长QTL,谷粒长宽比QTL与粒长和粒宽QTL,每穗粒数QTL与一次枝梗数和二次枝梗数QTL的关系。结果表明:鉴定出株高QTL的6个单片段代换系中有4个只检测出了主茎高QTL,其加性效应百分率为86.00%～99.55%,有1个只检测出了穗长QTL,其加性效应百分率为48.31%,有1个同时检测出了主茎高QTL和穗长QTL,其中主茎高QTL的加性效应百分率为81.72%,穗长QTL加性效应百分率为18.28%;在检测出主茎高QTL的7个单片段代换系中,有1个只检测出倒一节间长QTL,有2个只检测出倒二节间长QTL,有2个检测出倒一节间长QTL和倒二节间长QTL,有2个只检测出倒三节间长QTL;不同的单片段代换系中检测出的节间长的QTL加性效应百分率变化范围为-128.62%～172.07%;7个检测出谷粒长宽比QTL的单片段代换系中,有5个只检测出粒长QTL,1个只检测出粒宽QTL,1个同时检测出了粒长QTL和粒宽QTL;检测出每穗粒数QTL的3个单片段代换系中,有2个只检测出二次枝梗数的QTL,有1个同时检测出一次枝梗数QTL和二次枝梗数的QTL。这些结果表明,代换片段中如能检出复合性状QTL,也可以检出其构成性状QTL;复合性状QTL的加性效应的大部分可由其构成性状QTL的综合效应来解析,但相同的复合性状,不同代换片段检出的构成性状QTL不同。     

高含固率木质纤维素厌氧发酵物总DNA的4种提取方法比较

不同的样本特性和提取方法对获得微生物总DNA的质量有重要影响。文章基于高含固率木质纤维素厌氧发酵物腐殖酸、酚类物质含量高、质地均一性差、微生物浓度低的特点,研究了4种方法提取不同高含固率粪秸厌氧发酵物中微生物总DNA的效果。结果表明,常规的十二烷基磺酸钠法(sodium dodecyl sulfate,SDS)、十二烷基磺酸钠和溴化十六烷基三甲铵结合法(sodium dodecyl sulfate and cetyltrimethyl ammonium bromide,SDS-CTAB)和商业的粪便试剂盒法提取的DNA质量均较差,SDS法和试剂盒法未能获得聚合酶链式反应(polymerase chain reaction,PCR)扩增目的条带,SDS-CTAB法得到的条带较模糊;改进SDS-CTAB法获得的DNA杂质少、纯度高,具有较好的稳定性,A260/A280和A260/A230值分别为1.74~1.86和1.65~1.86,每克样品的DNA浓度在50 ng·μL^-1以上,电泳条带单一齐整、清晰明亮,PCR扩增的目的条带清晰度高,适宜后续分子生物学技术的分析。林格氏液洗脱、聚乙烯吡咯烷酮-40(Polyvinyl Pyrrolidone-40,PVP-40)洗涤液除杂以及裂解液和多种酶联合破壁是改进SDS-CTAB法获得该类专一性样本高质量微生物总DNA的关键步骤。     

Rapid screening of genetically modified ingredients in soybean and cotton processing by-product and waste using direct qPCR

To develop a simple and fast method for screening genetically modified ingredients from processing by-product and waste, direct quantitative PCR (qPCR) kit-Taqman which omitting multi genomic DNA preparing steps was developed in this study. A total of 18 oil crop processing by-products and wastes including 10 soybean and 8 cotton materials were collected from food processing factories. Compared with 2 commercial direct qPCR kits, conditions of DNA releasing procedure and PCR amplification were optimized. Element screening was performed at the initial step of genetically modified (GM) ingredient testing procedure via direct qPCR. GM event identification was carried out in positive samples by initial screening. Totally 5 screening elements (P–35S, T-NOS, Cp4-epsps, bar and pat) for soybean materials and 6 screening elements (P–35S, T-NOS, NPTII, Cry1Ac, bar and pat) for cotton samples were detected. In GM event identification, MON531 and MON1445 were found in cotton materials. Results were further confirmed by real-time PCR with DNA extraction and purification. The direct qPCR system proposed by this research was convenient for rapid screening and identification of GM ingredients in oil crop primary by-product and waste.     

Quantitative trait loci mapping for yield-related traits under low and high planting densities in maize (Zea mays)

Average maize yield per hectare has increased significantly because of the improvement in high-density tolerance, but little attention has been paid to the genetic mechanism of grain yield response to high planting density. Here, we used a population of 301 recombinant inbred lines (RILs) derived from the cross YE478 × 08–641 to detect quantitative trait loci (QTLs) for 16 yield-related traits under two planting densities (57,000 and 114,000 plants per ha) across four environments. These yield-related traits responded differently to high-density stress. A total of 110 QTLs were observed for these traits: 33 QTLs only under low planting density, 50 QTLs under high planting density and 27 QTLs across both densities. Only two major QTLs, qCD6 and qWKEL2-2, were identified across low- and high-density treatments. Seven environmentally stable QTLs were also observed containing qED6, qWKEL3, qRN3-3, qRN7-2, qRN9-2 and qRN10 across both densities, as well as qRN9-1 under low density. In addition, 16 and eight pairs of loci with epistasis interaction (EPI) were detected under low and high planting densities, respectively. Additionally, nine and 17 loci showed QTL × environment interaction (QEI) under low- and high-density conditions, respectively. These interactions are of lesser importance than the main QTL effects. We also observed 26 pleiotropic QTL clusters, and the hotspot region 3.08 concentrated nine QTLs, suggesting its great importance for maize yield. These findings suggested that multiple minor QTLs, loci with EPI and QEI, pleiotropy and the complex network of “crosstalk” among them for yield-related traits were greatly influenced by plant density, which increases our understanding of the genetic mechanism of yield-related traits for high-density tolerance.     

Mapping and validation of the quantitative trait loci for leaf stay‐green‐associated parameters in maize

Functional stay‐green is generally regarded as a desirable trait of varieties in major crops including maize. In this study, we used an F_3:4 recombinant inbred line population with 165 lines from a cross between a stay‐green inbred line (Zheng58) and a model inbred line (B73) using 211 polymorphic simple sequence repeat markers to map quantitative trait loci for three stay‐green‐associated parameters, chlorophyll content, photosystem II photochemical efficiency and stay‐green area, at maturity stage, detected a total of 23 quantitative trait loci (QTL) on nine chromosomes. Single QTL explained 3.7–13.5% of the phenotypic variance. Additionally, we validated some important stay‐green QTL using a heterogeneous inbred family approach and found that the stay‐green‐associated parameters were significantly correlated with the plant yield. This study may contribute to a better insight into the regulatory mechanism behind leaf stay‐green in maize and a novel development of elite maize varieties with delayed leaf senescence through molecular marker‐assisted selection.     

菠萝酸性转化酶基因家族初步生物信息学及表达分析

酸性转化酶（acid invertase, AIN）在菠萝采后蔗糖降解过程中起着重要作用,基于菠萝全基因组数据库,预测菠萝AIN家族基因并进行生物信息学分析,解析其在采后菠萝不同贮藏温度下的表达变化情况,为阐明AIN基因在采后菠萝果实贮藏特性中的作用奠定基础。以水稻AIN家族基因为探针,在菠萝全基因组中鉴定到2个菠萝细胞壁酸性转化酶基因（cell wall acid invertase, CWIN）和2个液泡酸性转化酶基因（vacuolar acid invertase, VIN）,分别命名为AcCWIN1、AcCWIN2、AcVIN1、AcVIN2,设计编码区引物进行测序验证,并进行生物信息学分析。进化分析结果表明,AcCWIN1、AcCWIN2和AcVIN1、AcVIN2蛋白分别归于细胞壁酸性转化酶和液泡酸性转化酶2个进化支上,且均属于糖基水解酶家族GH32,基因结构、保守域和保守基序均一致。荧光定量分析结果表明,菠萝果肉中AcVIN1和AcVIN2在果实采后贮藏过程中表达量升高,且AcVIN1在发生黑心病的部位大量表达,而AcCWIN1和AcCWIN2在采后贮藏过程中表达量逐渐降低,且随着贮藏温度的升高其表达量降低,预示AcVIN1、AcVIN2较AcCWIN1、AcCWIN2在菠萝采后蔗糖降解和黑心病的发生方面发挥着更为重要的作用。