牛角瓜根的抑菌活性研究

以牛角瓜根为实验材料,采用索氏提取法,以4种不同溶剂(石油醚、氯仿、乙酸乙酯、甲醇)对牛角瓜的根进行提取,并采用纸片琼脂扩散法对粗提物的抑菌活性进行了评价,供试菌包括白色念珠菌等6种真菌和金黄色葡萄球菌等7种细菌。结果表明:石油醚粗提物对枯草芽孢杆菌、白色念珠菌、黑曲霉菌、大肠埃希氏菌都表现出良好的抑菌活性,且抑菌效果接近或优于阳性对照氟康唑,对尖孢镰刀菌和鲍曼不动杆菌也表现出良好的抑菌活性;氯仿粗提物对白色念珠菌及粪链球菌表现出良好的抑菌活性;乙酸乙酯粗提物对粪链球菌表现出最好的抑菌活性;甲醇粗提物的抑菌活性较弱。     

Investigation of biofilm production and icaA and icaD genes in Staphylococcus aureus isolated from heifers and cows with mastitis

Biofilm formation and antimicrobial resistance of Staphylococcus aureus are important virulence factors in cases of mastitis in dairy cows. However, few studies have investigated mastitis strains isolated from heifers. Within this context, the objective of the present study was to investigate biofilm formation on Congo red agar, the presence of the icaA and icaD genes by polymerase chain reaction (PCR), and the percentage of in vitro antimicrobial resistance of 110 S. aureus isolates from mammary gland secretions of heifers and cows with mastitis. PCR detected the icaA and icaD genes in 98% and 100% of isolates, respectively. However, only 55.5% of all isolates produced a biofilm on Congo red agar. Antimicrobial susceptibility testing revealed that 47.0% of isolates from heifers and 70.4% of isolates from cows were resistant to at least one of the antimicrobial agents tested. Resistance to penicillin and/or ampicillin was the most frequent (44.5%). These results indicate the need to implement prophylactic and control measures of mastitis for heifers. Heifers and cows can carry resistant strains with the capacity of biofilm production, a fact representing a threat to public health and animal well‐being and generating losses to dairy farmers.     

Phenotypic and genotypic characterization of canine pyoderma isolates of Staphylococcus pseudintermedius for biofilm formation

Biofilm-forming ability is increasingly being recognized as an important virulence factor in several Staphylococcus species. This study evaluated the biofilm-forming ability of sixty canine derived clinical isolates of S. pseudintermedius, using three phenotypic methods, microtiter plate test (MtP), Congo red agar method (CRA) and tube adherence test, and the presence and impact of biofilm-associated genes (icaA and icaD). The results showed that icaA and icaD genes were detected concomitantly in 55 (91.7%) of 60 isolates. A majority (88.3%) of the strains screened had matching results by the tube adherence test, MtP and PCR analysis. Better agreement (95%) was found between the PCR-based analysis and the CRA. Results of the icaA and icaD gene PCRs showed good agreement with CRA results, with a kappa of 0.7. Comparing the phenotypic methods, the statistical analysis showed that the agreement among the phenotypical tests using categorical data was generally good. Considering two classes (biofilm producer and biofilm non-producer), the percentage of matching results between the CRA method and the tube adherence test and between the CRA method and the MtP was 93.3%. A concordance of 100% was revealed between the MtP and the tube adherence test. The results indicate a high prevalence of the ica genes within S. pseudintermedius isolates, and their presence is associated with in vitro formation of a biofilm. A combination of phenotypic and genotypic tests is recommended for investigating biofilm formation in S. pseudintermedius.     

牛源多杀性巴氏杆菌主要荚膜群和脂多糖基因型两组三重PCR检测方法

为掌握国内牛源多杀性巴氏杆菌流行的主要荚膜群及脂多糖基因型,建立了检测多杀性巴氏杆菌(Pm)及其A、B荚膜群的三重PCR(PmAB-3PCR)以及检测3个脂多糖基因型的三重PCR(LPS-3PCR)方法,检验了其特异性和敏感性,用感染Pm小鼠的组织和人工污染Pm的牛鼻拭子样品进行了临床模拟检验,并对30株P m进行了PCR检测和传统血清学分型比较。结果显示:PmAB-3PCR特异性好,PmAB-3PCR和LPS-3PCR的DNA检测限分别为10~100 pg和1 ng,二者对菌液的检测限分别为200~2000 CFU和2000~20000 CFU。模拟临床样品PmAB-3PCR检测结果与细菌分离结果100%一致。PmAB-3PCR与琼扩试验的符合率为84%,二者检出的阳性率分别为88%和72%,差异不显著(P>0.05);LPS-3PCR与琼扩试验的符合率为60%,检出的阳性率分别为90%和50%,差异显著(P<0.05)。结果表明,建立的两组三重PCR方法准确高效且重复性好,可取代常规血清学方法用于国内牛巴氏杆菌病的诊断、流调和疫苗株筛选。     

不同培养方法及培养密度对绵羊腔前卵泡发育的影响

通过对绵羊腔前卵泡培养方法的研究,能够探索出适宜绵羊腔前卵泡生长发育的培养条件,以及为进一步揭示绵羊卵泡生长发育的规律奠定一些基础。试验一通过使用微滴法和琼脂培养基法培养腔前卵泡,观察其对绵羊腔前卵泡生长的影响;试验二通过绵羊腔前卵泡单独培养与群体培养,探讨不同培养密度对绵羊腔前卵泡生长的影响。试验结果显示,使用微滴法与琼脂培养基法培养的绵羊腔前卵泡,在培养3天时,卵泡直径增加值和存活率差异均不显著(P0.05);但培养6天时,琼脂培养基法培养的腔前卵泡的直径增加值和存活率均极显著高于微滴法培养的腔前卵泡(P0.01);在使用琼脂培养基培养的条件下,对绵羊腔前卵泡进行单独培养和群体培养,在培养的6天时间内二者卵泡直径增加值和存活率差异均不显著(P0.05)。试验结果表明,琼脂培养基法是较为适宜的绵羊腔前卵泡体外培养方法;培养密度对绵羊腔前卵泡体外培养并无显著影响。     

琼脂及琼脂糖中3,6-内醚半乳糖的测定

本文研究了间苯二酚法对琼脂及琼脂糖样品中的3,6-内醚半乳糖的测定方法。实验中,根据间苯二酚试剂与3,6-内醚半乳糖和果糖的浓度在0.25μmol/mL以下时,具有近乎相同的标准比色曲线的原理,分别对测定波长、显色温度、显色时间和比色时间等检测条件进行优化。结果显示,用间苯二酚法测定3,6-内醚半乳糖的最佳检测条件为:测定波长554 nm、显色温度80°C、显色时间15 min;在该条件下,对琼脂及琼脂糖样品中的3,6-内醚半乳糖进行测定,方法的相对偏差为0.94%～1.27%,加标回收率为105.4%～111.04%。表明该方法操作简便,具有较高的准确性和重复性。     

基于大豆膳食纤维的高黏度样品细菌总数检测方法研究

[目的]研究基于大豆膳食纤维的高黏度样品的细菌总数检测方法。[方法]对大豆膳食纤维样品的前处理过程和培养基选择进行了研究。[结果]在大豆膳食纤维样品的前处理过程中,与普通方法相比,采用称重法和RAININ Pos-D外置活塞式微量移液器量取稀释液所测得的细菌总数较高。3种计数培养基中,采用3M PetriflimTM菌落总数测试片的菌落总数测定结果最好,其次是采用平板计数琼脂,最差是采用TTC营养琼脂;3M测试片和TTC营养琼脂的测定结果的精密度较好、人员计数偏差较小,而平板计数琼脂相对较差。[结论]为检测具有颗粒、高吸水性及高黏度等性质的样品提供了参考。     

一种大豆疫霉菌(Phytophthora sojae)纯营养菌丝的培养方法

比较了胡萝卜琼脂和玻璃纸胡萝卜琼脂培养大豆疫霉菌(Phytophthora sojae)对菌丝生长速度、卵孢子产生、孢子囊产生时间及RNA提取效果的影响。结果表明,玻璃纸胡萝卜琼脂培养的大豆疫霉菌,10 d内不会产生卵孢子,当大豆疫霉菌从玻璃纸上取下时,不会带有培养基,能够提取出高质量的RNA,满足差异显示反转录PCR对RNA质量的要求,是一种适合大豆疫霉菌纯营养菌丝的培养方法。     

Seaweed Hydrocolloid Production: An Update on Enzyme Assisted Extraction and Modification Technologies

Agar, alginate, and carrageenans are high-value seaweed hydrocolloids, which are used as gelation and thickening agents in different food, pharmaceutical, and biotechnological applications. The annual global production of these hydrocolloids has recently reached 100,000 tons with a gross market value just above US$ 1.1 billion. The techno-functional properties of the seaweed polysaccharides depend strictly on their unique structural make-up, notably degree and position of sulfation and presence of anhydro-bridges. Classical extraction techniques include hot alkali treatments, but recent research has shown promising results with enzymes. Current methods mainly involve use of commercially available enzyme mixtures developed for terrestrial plant material processing. Application of seaweed polysaccharide targeted enzymes allows for selective extraction at mild conditions as well as tailor-made modifications of the hydrocolloids to obtain specific functionalities. This review provides an update of the detailed structural features of κ-, ι-, λ-carrageenans, agars, and alginate, and a thorough discussion of enzyme assisted extraction and processing techniques for these hydrocolloids.