Nested PCR for the rapid detection of TB from pleural fluid at HUKM Malaysia

The aim of the present study is rapid detection of tuberculosis from pleural effusion of suspected patients. Molecular technique Nested Polymerase Chain Reaction (PCR) was used for the purpose. A total of 67 pleural fluid collected at Hospital University Kebangsaan Malaysia during May 2005 to October 2006 were sent to Microbiology Laboratory enrolled in the study. Detection rate of Mycobacterium tuberculosis in pleural effusion was 0% by acid-fast bacilli (AFB) staining and 1.5% by culture on Lowenstein-Jensen medium. Mycobacterium tuberculosis was detected by PCR in 9% of the cases. PCR of pleural fluid had 19% sensitivity and 96% specificity, compared to AFB staining (0% sensitivity and 100% specificity) and culture (4% sensitivity and 100% specificity). PCR also has 67% Positive Predictive Value (PPV) and 72% Negative Predictive Value (NPV) in detecting Mycobacterium tuberculosis. Culture ofpleural fluid has 100% PPV and 71% NPV while AFB staining has 0% PPV and 31% NPV. This preliminary study showed that PCR is a rapid method for detection of Mycobacterium tuberculosis in pleural fluid but its sensitivity is not up the marked.     

Detection of Microsatellite Instability by High-Resolution Melting Analysis in Colorectal Cancer

Background: Colorectal cancer is the third most common cancer worldwide. MSI is a molecular marker of a deficient MMR system and happens in almost 15% of CRCs. Because of a wide frequency of MSI⁺ CRC in Iran compared to other parts of the world, the importance of screening for this type of cancer is highlighted. Methods: The most common MSI detection technique is a fluorescent PCR-based method in which fragments are analyzed by CE. This technique is very time-consuming, difficult, and expensive. We sought to develop and evaluate a proper method with high accuracy, specificity, and sensitivity to screen the MSI⁺ CRC. A HRM analysis procedure is relying on the analysis of the melting curve attributes. Low cost, feasibility, high specificity, and sensitivity are outstanding attributes of HRM analysis. Results: Five mononucleotide microsatellite markers, including BAT-25, BAT-26, NR-21, NR-24, and NR-27, in 25 archival CRC tumor tissue samples were compared with normal tissue adjacent using HRM method. The specificity and sensitivity of BAT-25 with HRM method were 100% compared to CE, while other markers had lower sensitivity. However, when all the markers were considered together, the sensitivity and specificity became 100%. The number of MSI⁺ samples was 56%, which shows a higher ratio than previous Iranian studies. The highest MSI was related to BAT-26 (52%). Conclusion: The HRM method is much simpler and more cost-effective than current MSI techniques, and its sensitivity and accuracy are comparable. Therefore, it can serve as an alternative method in cases where CE is unavailable. Key Words: Biomarkers, Microsatellite instability, Colorectal cancer     

槟榔隐症病毒1型TaqMan实时荧光定量PCR检测方法的建立

为了建立一种快速、精准且能定量分析槟榔隐症病毒1型（Areca palm velarivirus 1, APV1）的检测方法。本研究参照GenBank中已登录的APV1全基因组序列,在p21蛋白基因保守区域和p60蛋白基因保守区域之间设计1对特异性RT-PCR引物,并构建阳性重组质粒;在p60蛋白基因保守区域部分设计荧光定量PCR特异性引物和TaqMan-MGB探针,利用阳性重组质粒,构建标准曲线,并对该检测方法的敏感性、特异性、稳定性及其应用效果进行验证。结果显示：该方法对阳性质粒标准品检测的敏感性达3.0×10¹ copies/μL级别,是常规RT-PCR敏感性的100倍;标准曲线显示,C_t值与拷贝数的对数呈线性关系,其标准曲线方程为y=-3.2748x+42.957,扩增效率为102%,相关系数R²为0.9992;该方法对APV1的检测具有较好的特异性,其他槟榔病原物及内生菌不会对本方法产生非特异性干扰;批内与批间重复性试验结果显示,该方法对APV1的检测具有较好的重复性和稳定性;利用该方法对海南万宁、琼海、定安、文昌等4个市（县）采集的181份疑似样品进行检测,阳性率分别为91.95%、86.95%、89.65%、73.68%,其阳性的病毒拷贝数最高达1.59×10⁷copies/μL,表明海南部分地区APV1病毒载量较高、流行情况比较严重。     

Serum Krebs von den Lungen-6 for Predicting the Severity of COVID-19 Lung Injury: A Systematic Review and Meta-Analysis

Introduction:Lung injury is common in COVID-19 patients. The severity of lung injury appears to be reflected in serum KL-6, a glycoprotein expressed on type II alveolar epithelium. This study aims to assess the role of serum KL-6 in reflecting the severity of lung injury in COVID-19 patients. Methods:A systematic search was conducted in Scopus, PubMed, Wiley Online Library, and ProQuest. Articles were screened based on several eligibility criteria and assessed for study quality using NOS. Results:This systematic review included four studies involving a total of 151 adult COVID-19 patients. Pooled analysis revealed that serum KL-6 was significantly higher in severe patients (SMD = 1.16; 95% CI = 0.69–1.63) with moderately high pooled sensitivity (79%; 95% CI = 61–91%) and specificity (86%; 95% CI = 72–95%). Conclusion:High serum KL-6 may depict more severe lung injury in COVID-19 patients with moderately high sensitivity and specificity. Key Words: Biomarker, COVID-19, Infectious disease, Krebs von den Lungen-6, Lung injury     

Comparative efficacy sensitivity and specificity of the tests used for the Diagnosis of Helicobacter pylori

Abstract: A total number of 157 samples were examined by 4 different tests-In-house rapid urease (iRUT), Culture, Histopathology and Immunochromatography (Immuno CardSTAT) for the detection of Helicobacter pylori from the patients reported to Hospital Kuala Lumpur, Malaysia during 2007 to 2008. Out of the samples examined 47 (29.9%) were positive for H. pylori by the tests used in the laboratory. Efficacy of detection of the bacteria by the tests- In-house rapid urease, Culture, Histopathology and Immuno CardSTAT were 31.8, 13.9, 30.3 and 32.8%, respectively. However, sensitivity and specificity of the iRUT were 91.5 and 93.6%, respectively and the Positive Predictive Value (PPV) was 86% and the Negative Predictive Value (NPV) was 96.3%. The sensitivity for Immuno CardSTAT rapid test was 100% and the specificity was 79.3%. The PPV was 50% and the NPV was 100%. Convenient methods to the authors were 'In house rapid urease test and Immunochromatography though variability of specificities were observed.     

橡胶树炭疽菌生物学和侵染特征研究

橡胶树炭疽病是危害我国橡胶树的一种重要叶部病害,致病菌为胶抱炭疽菌。对其生物学特性研究结果表明,在28 ℃下,有利于菌丝体生长、产孢和孢子萌发 。最适合菌丝体生长的pH值为6。黑暗下有利于孢子萌发和菌丝体生长,但产孢则不如在光照条件下。在100 %相对湿度下孢子萌发率最高。在葡萄糖、橡胶叶汁和琼脂胶等营养液中, 孢子萌发率高于清水对照。对其侵染特征研究结果表明,主要侵染源为受害嫩梢和半枯死枝条,寄主范围广泛。病菌可从伤口、气孔和表皮3种途径入侵,风雨是传播媒介。潜育期一般3~6d,条件最适宜时潜育期为1~ 2d。田间气温21~24℃、相对湿度大于95%时,病菌产孢多,侵人迅速,病斑扩展快     

Prediction of lactic acid bacterial numbers on lucerne

The numbers of lactic acid bacteria (LAB), as measured by Rogosa SL agar, on lucerne ( Medicago sativa ) chopped for ensiling were predicted using two methods. The first prediction method consisted of regression equations developed from two previous harvesting seasons and required inputs of wilting time between mowing and chopping, average wilting temperature and moisture content of the lucerne at chopping. The second method simulated the growth of LAB on the lucerne during wilting and needed similar inputs. The former method predicted LAB counts within one order of magnitude more than 95% of the time except for lucerne samples drier than 60% dry matter. The prediction of the simulation model had more variation than those of the regression equations, but the simulation model was applicable over a wider range of conditions. These results, together with earlier studies, provide a preliminary basis for determining when a silage inoculant containing LAB will be most beneficial.     

A fatty acid glycoside from a marine-derived fungus isolated from mangrove plant Scyphiphora hydrophyllacea

To study the antimicrobial components from the endophytic fungus A1 of mangrove plant Scyphiphora hydrophyllacea Gaertn. F., a new fatty acid glucoside was isolated by column chromatography from the broth of A1, and its structure was identified as R-3-hydroxyundecanoic acid methylester-3-O-α-l-rhamnopyranoside (1) by spectroscopic methods including 1D and 2D NMR (HMQC, (1)H-(1)H COSY and HMBC) and chemical methods. Antimicrobial assay showed compound 1 possessed modest inhibitory effect on Saphylococcus aureus and methicillin-resistant S. aureus (MRSA) using the filter paper disc agar diffusion method.     

水稻GA20ox-2基因mRNA的TaqMan荧光定量RT-PCR检测

 成功建立了一项基于TaqMan 实时荧光定量的RT-PCR技术,定量分析水稻半矮化关键基因之一GA20ox-2转录水平。该技术体系中重组质粒标准品的制备方法具有很好的实用性;质粒标准品对基因GA20ox-2表达的实时定量准确、可靠、便捷。标准曲线表明,所建立的GA20ox-2基因mRNA表达实时荧光定量PCR检测方法,特异性好,灵敏度高,可达102拷贝;线性范围广,可达102～107拷贝;扩增效率高（E=100.3%）;稳定性、重复性好,可靠性高,批内和批间变异系数仅分别为0.12%～0.31%和0.21%～0.34%;循环阈值与PCR 体系中起始模板量的对数值之间有着良好的线性关系（^r＝0.999）,可对GA20ox 2基因表达进行准确实时定量。     

NCM-ELISA检测马铃薯Y病毒(PVY)技术的研究及应用

血清学方法是病毒检测的主要手段。本试验通过对马铃薯Y病毒(PVY)的提纯,免疫家兔制备PVY抗血清,并提取PYV免疫球蛋白IgG作为NCM-ELISA反应为一抗,以市售羊抗兔抗血清为二抗,在硝酸纤维素膜(NCM)上进行NCM-ELISA反应检测PVY,建立PVY NCM-ELISA检测反应体系。试验结果显示,NCM-ELISA具有反应特异性强,灵敏度高的优点,检测植物叶片样品的最高稀释度可达到1:250。通过对田间40份样品的NCM-ELISA和DAS-ELISA检测比较,其检测结果吻合率达到100%。由于NCM-ELISA方法可以将样品点在硝酸纤维素膜上,并且可贮存几个星期或将膜送到其他实验室进行检测,因此具有操作简单,使用方便,检测成本低等优点。     

不同剂型草甘膦对金龟子绿僵菌MA4菌株生长发育的影响

测试3种不同的草甘膦剂型在不同浓度下对MA4菌株孢子萌发、菌落生长、产孢量的影响。结果表明,飞达红88.8％草甘膦铵盐可溶性粒剂对孢子萌发与菌落生长抑制作用最强,10％草甘膦水剂对产孢的抑制作用最大,95％的氨盐影响均相对较小。同时研究发现,草甘膦各剂型中的不同助剂对金龟子绿僵菌菌株的生长与产孢的影响较大。     

野生、栽培大豆愈伤组织耐盐性与植株盐胁迫下丙二醛(MDA)含量的关系

以野生、栽培大豆共14份材料的下胚轴作为外植体诱导愈伤组织，在含不同浓度NaCl的培养基上鉴定愈伤组织的耐盐性，NaCl浓度(w／v)分别为0％(CK)，0．1％，0．2％，0．3％，0．4％，0．5％，0．6％，0．7％，0．8％；14份材料的幼苗在NaCl为0．3％的盐土环境中生长25d后，测其幼苗的丙二醛(MDA)含量。试验结果表明：愈伤组织水平上，栽培大豆品种中黄4、通农5、吉林35耐盐性较强，不同材料之间愈伤组织的耐盐性有明显差异，耐盐性强的可在0．8％NaCl浓度下生长，耐盐性弱的只能忍受0．4％NaCl浓度；本试验确定大豆愈伤组织耐盐性鉴定适宜的浓度为0．6％。在盐胁迫下，植株MDA含量在0．3％NaCl条件下均比在CK土壤中高，但随胁迫时间的延长而呈现不同的变化趋势。综合大豆植株MDA含量的变化和愈伤组织的耐盐性之间，并未发现相关性。     

“热研11号”黑籽雀稗植株再生体系的建立

以热带牧草"热研11号"黑籽雀稗(Paspalum atratum cv.Reyan No.11)种子为材料,对其外植体植株的再生过程进行系统研究.结果表明,以MS无机盐 9.0mg/L维生素B1 9.5mg/L维生素B6 4.5mg/L尼克酸 1.0mg/L水解酪蛋白 30.0g/L蔗糖 8.0g/L琼脂为基本成分(MSM),附加植物激素类物质2.0mg/L2,4-D时,适合种子的愈伤组织形成,愈伤诱导率可达65%:继代培养基附加1.0mg/L2,4-D和0.1mg/LKT:分化的培养基附加6.0mg/L6-BA,分化率可达50%;生根培养基附加0.5mg/L激素类物质NAA,生根率100%.完成植株再生约需13周.     

进口大豆中菜豆晕疫病菌巢式PCR检测方法的建立

为实现对进口大豆样品中菜豆晕疫病菌的快速检测,本研究根据菜豆晕疫病菌的argK特异性序列设计引物52B/8F和24B/24F,建立了巢式PCR检测方法,并进行特异性和灵敏度验证。试验结果表明,利用此检测方法对多种参试菌株进行检测时,只有菜豆晕疫病菌呈阳性反应,而其他病菌不产生扩增反应;巢式PCR检测方法对菜豆晕疫病菌基因组DNA和菌悬液检测时,其灵敏度分别达到0.916×10-4ng/μL和2.4×103CFU/m L,比常规PCR灵敏度高1 000倍。在对100份进口大豆样品检测时,3份样品扩增到了特异性条带,测序分析结果证明3份大豆样品中携带的病菌确实为菜豆晕疫病菌。本研究所建立的巢式PCR检测方法具有特异性强、灵敏度高、检测时间短并且检测准确率高等特点,可用于口岸菜豆晕疫病菌的检测。     

New Isocoumarin Derivatives and Meroterpenoids from the Marine Sponge-Associated Fungus Aspergillus similanensis sp. nov. KUFA 0013

Two new isocoumarin derivatives, including a new 5-hydroxy-8-methyl-2H, 6H-pyrano[3,4-g]chromen-2,6-dione (1) and 6,8-dihydroxy-3,7-dimethylisocoumarin (2b), a new chevalone derivative, named chevalone E (3), and a new natural product pyripyropene S (6) were isolated together with 6, 8-dihydroxy-3-methylisocoumarin (2a), reticulol (2c), p-hydroxybenzaldehyde, chevalone B, chevalone C, S14-95 (4), and pyripyropene E (5) from the ethyl acetate extract of the undescribed marine sponge-associated fungus Aspergillus similanensis KUFA 0013. The structures of the new compounds were established based on 1D and 2D NMR spectral analysis, and in the case of compound 3, X-ray analysis was used to confirm its structure and the absolute configuration of its stereogenic carbons. Compounds 1, 2a–c and 3–6 were evaluated for their antimicrobial activity against Gram-positive and Gram-negative bacteria, Candida albicans ATCC 10231, and multidrug-resistant isolates from the environment. Chevalone E (3) was found to show synergism with the antibiotic oxacillin against methicillin-resistant Staphylococcus aureus (MRSA).     

NAA对盐胁迫下水稻根系生长的缓解效应

采用水培的方法,研究了外源NAA对盐胁迫下水稻幼苗根系生长的影响.结果 表明,20~100 mg· L-1 NAA处理均能缓解100 mmol·L-1 NaCl胁迫对水稻幼苗根生长的抑制效应.NAA对根系生长的缓解效应随处理浓度的增加呈现先增后降的趋势,其中以80 mg·L-1NAA的效果最明显.NAA可通过抑制盐胁迫下水稻根尖活性氧积累和降低根尖细胞质膜透性来保护根系正常的生理活性.     

水稻不同品种耐盐限度研究初报

了解水稻不同品种的耐盐限度,对于盐渍土区稻作生产的持续、稳定发展有着重要意义。试验表明,土壤全盐5.94g/kg时,盐粳68、花粳8、辽粳294和盐粳34的发芽率均不低于90%。花粳15分蘖期最高耐盐为14g/kg,盐粳68拔节孕穗期最高耐盐为17g/kg,盐丰47-6成熟期最高耐盐为12g/kg。12个供试水稻品种,其中盐粳68、花粳8、辽粳9和92-71耐盐能力强;花粳15、抗盐100、盐丰47-6、辽粳294、盐粳34和盐粳31耐盐能力次之。     

油菜与诸葛菜属间远缘杂交的初步研究

为了克服甘蓝型油菜与诸葛菜远缘杂交双受精前雌雄不亲和性,本实验采用60ppm的赤霉素、100ppm的甘氨酸涂抹油菜柱头、10ppm的H_3BO_3 5％的蔗糖涂抹去掉柱头的油菜花柱和蒙导法。赤霉素效果最好,获10粒种子,甘氨酸有一定效果得到3粒种子。而截去柱头和蒙导法均未见效果。在获得的13粒发育不全种子中,7粒种子萌发成苗。幼苗形态介于油菜与诸葛菜之间。     

Antimicrobial susceptibility pattern of microorganisms involved in the pathogenesis of surgical site infection (SSI); A 1 year of surveillance

The aim of this study is to identify the antibiotic sensitivity pattern of pathogens involved in the process of surgical site infection, in surgical wards. Changes made in the pattern of antibiotic use will result in different microorganism susceptibility patterns, which needs correct determination for precise empiric antibiotic therapy. One thousand patients (62% men and 38% women, 18- 74-years-old, with mean age 43 +/- 8)) who underwent surgical treatment, in Alzahra University Hospital, Isfahan University of Medicine, Isfahan, Iran, were studied from 2005 to 2006. Surgical wound infections, based on the reported criteria, were aspirated for culturing within 1 plus gram staining of prepared smears. Minimum Inhibitory Concentrations (MICs) were determined for samples and all derived data were compared by SPSS 13 and WHO net 5 software. The prevalence of SSI was 13.3% with 150 positive cultures, totally. Of 150 bacteria, isolated from surgical site infections Staphylococcus aureus had most frequency (43%). Resistance of isolated organisms was 41.7% in amikacin, 65 and 78.6% in ceftazidime, 85.7% in ceftriaxone, 61.5% in ciprofloxacin, 78.8% in gentamicine, 6.4% in imipenem, 13% in meropenem and 70.6% in trimethoprim/sulfamethoxazole, respectively. 78.9% of Staphylococcus aureus isolates were MRSA and vancomycine was the most effective antibiotic without any resistance. Among 10 isolates of coagulase negative Staphylococcus, no vancomycine resistance was seen, but in contrast all cases were resistant to oxacillin. The most common gram negative organism was Klebsiella (18 isolates) in which 100 and 80% were sensitive to imipenem and meropenem, respectively. Seventeen cases were E. coli, in which the most sensitivity was to meropenem (80%) and imipenem (77.8%). Thirteen cases of Pseudomonas were detected, in which 16.7% were resistant to imipenem and 8.3% to meropenem. Our results demonstrated that the total antibiotic resistance is increasing among SSIs, with an up sloping pattern, which will contact with a constant empiric antibiotic therapy. So, precise up to date antibiogram tantalize us toward balancing the rate of total antibiotic resistance to SSIs.