chapter實驗室的工具研究微生物的方法PPT課件

1、Main Themes Methods of Culturing（培養(yǎng)） Microorganisms（微生物） The Microscope（顯微鏡）第1頁/共82頁 Over the past 30 years,nearly 20 new infectious（傳染性的） agents（介質(zhì)） have appeared suddenly and unexpectedly（意外的）.第2頁/共82頁Chapter Overview A driving force of microbiology has been to find ways to visualize（顯現(xiàn)） and handl

2、e（處置 ）microorganisms; Microbes are managed and characterized （概括特征）by implementing the Five I,S; The microscope is a powerful tool for magnifying（放大） and resolving cells and their parts;第3頁/共82頁Methods of Culturing Microorganisms Problems: Most habitats（棲息地） harbor（居?。?microbes in complex（復雜的） assoc

3、iations ,so it is often necessary to separate the species from one another; To maintain and keep track of （了解）such small reserarch subjects,mircrobiologists（微生物學家） usually have to grow them under artificial conditions; They are invisible and widely distributes（分布廣泛）,and undesirable（意外的） ones can be

4、introduced into an experiment and cause misleading results.第4頁/共82頁 If you want to study a specified（特定地） microorganism ,you should do experiments step by setp -according to the Five I,S.第5頁/共82頁第6頁/共82頁第7頁/共82頁INOCULATION:PRODUCING A CULTURE To cultivate（培養(yǎng)） microorganisms ,one introduces（引入） a tin

5、y sample into a container（容器） of nutrient（營養(yǎng)物） medium,which provides an environment in which they multiply（繁殖）.This process is called inoculation(接種).第8頁/共82頁ISOLATION:SEPARATE ONE SPECIES FROM ANOTHER If an individual bacterial cell is separated from other cells and provided adequate（充足的） space on

6、a nutrient surface,it will grow into a discrete（分立的） mound（堆積物） of cells called a colony(菌落).第9頁/共82頁Isolation Technique第10頁/共82頁Methods for isolating bacteria第11頁/共82頁The spreading tool第12頁/共82頁MEDIA:PROVIDING NUTRIENTS IN THE LABORATORYA medium（培養(yǎng)基） is a liquid or gel（凝膠） designed to support the g

7、rowth of microorganisms or cells .第13頁/共82頁Types of Media Physical form Chemical composition（組成） Functional type(table 3.1)第14頁/共82頁Physical States of Media Liquid media are defined as water-based that do not solidify（固化） at temperatures above freezing and that tend to flow freely when the container

8、 is tilted（傾斜）.第15頁/共82頁Sample liquid media第16頁/共82頁Semilolid media They contain some an amount of solidifying agent that thickens them but does not produce a firm substrate（基礎）.第17頁/共82頁Sample semisolid media第18頁/共82頁Solid media Solid media provide a firm surface on which cells can form discrete co

9、lonies and are advantageous（適合的） for isolating and culturing bacteria and fungi（真菌）.第19頁/共82頁Solid media第20頁/共82頁Agar(瓊脂) Isolated from the red alga Gelidium（石花菜屬 ）; It is solid at room temperature; Onced liquefied（液化）,agar does not resolidify until it cools to 42； Semisolid media(0.3-0.5%)、solid me

10、dia (0.5-2%).第21頁/共82頁 Nutrient agar（營養(yǎng)瓊脂） is a microbiological growth medium commonly used for the routine（常規(guī)的） cultivation of non-fastidious（苛求的） bacteria.第22頁/共82頁 Nutrient agar typically contains (w/v): 0.5% peptone （蛋白胨） 0.3% beef extract （牛肉膏） 1.5% agar （瓊脂） pH adjusted to neutral at 25 . Nutr

11、ient broth does not contain agar.第23頁/共82頁Peptone （蛋白胨）Any water soluble（可溶的） product, a mixture of polypeptides（多肽） and amino acids（氨基酸） formed by the partial hydrolysis（水解作用） of protein（蛋白質(zhì)）.第24頁/共82頁Peptone（蛋白胨）第25頁/共82頁Chemical Content of Media Chemical content of media are various(多種多樣的); They

12、are classified into synthetic（合成的） ingredients and non-synthetic（非合成的） ingredients（成分）; Natural media and synthetic media;第26頁/共82頁Natural comounds(天然化合物)Peptone（蛋白胨）Beef extract（牛肉膏）Potato（土豆）Bread（面包）第27頁/共82頁Synthetic compounds(合成化合物)NaclkH2PO4Mgso4第28頁/共82頁MediaUndefined media(天然培養(yǎng)基) all natural

13、 resourcesChemical defined media all chemical compounds Semidefined media(半組合培養(yǎng)基) natural resources +chemical compounds第29頁/共82頁Media to Suit Every Functions Microbiologists have many types of media at their disposal(實驗); Media are used for primary isolation,to maintain cultures in the lab,to detemi

14、ne chemical and growth characteristics（特征） ,and for other functions; General media（基礎培養(yǎng)基）、enriched meida（加富培養(yǎng)基）、selective and differential media（選擇分化培養(yǎng)基）、第30頁/共82頁Basal(general) media Basal media(基礎培養(yǎng)基) are designed to grow as broad a spectrum(范圍) of microbes as possible. Peptone （蛋白胨） Beef extract（

15、牛肉膏） Nacl（氯化鈉） Agar（瓊脂） Water （水）第31頁/共82頁Enriched media(加富培養(yǎng)基) An enriched media contains complex(復雜的) orgainc（有機物） substances such as blood ,serum（血清），hemoglbin （血色素）,or special growth factors that certain species must have in order to grow.第32頁/共82頁 When a media is prepared which have a nutrient

16、constituent（成份） which encourage the growth of particular bacteria or microbe is called enriched media, such as blood agar（血瓊脂） and chocolate agar(巧克力瓊脂). 第33頁/共82頁Examples of enriched media第34頁/共82頁Selective Media(選擇性培養(yǎng)基) A selective media contains one or more agents that inhibit(禁止) the growth of A

17、 microbe but not B and therby encourages,or selcets B microbe and allow it to grow.第35頁/共82頁Comparison of Selective media and general media第36頁/共82頁Example of selective media第37頁/共82頁Differential media(鑒別培養(yǎng)基) Differential media grow several types of microorganisms and are desigened to display（展示） vi

18、sible（可見的） differences among those microorganisms.第38頁/共82頁EMB meida（伊紅美藍瓊脂培養(yǎng)基）第39頁/共82頁Media that differentiate characteristics第40頁/共82頁A reducing medium contains a substance that absorbs oxygen（氧氣）Of slowing the penetration（滲透） of oxygen in a medium.Transport （運輸） mediaare used to maintain and pre

19、serve specimens that have to be held for a period of time before analysis.第41頁/共82頁Carbohydrate fermentation in broths（肉湯）第42頁/共82頁INCUBATION,INSPECTION,AND IDENTIFECATION The usual temperatures used in lab fall between 20 C and 40 C.第43頁/共82頁 If you incubate in the liquid meida,you may see cloudine

20、ss（混濁 ）,sediment （沉淀）and colour. A common manifestation(例證) of growth on solid media is the appearance of colonies（菌落）.第44頁/共82頁Liquid media第45頁/共82頁Various conditions of cultures第46頁/共82頁 A mixed culture is a container that holds two or more identified,easily differentiated（不同的） species of microorg

21、anism.第47頁/共82頁InspectionIdentificationBiochemistry（生化） and molecular tests第48頁/共82頁Maintenance and disposal of Cultures Disposed microorgansimsSterilizationBurningBoiling第49頁/共82頁The Microscope:Window on an Invisible realmMicroscopeMagnificationShowing detail第50頁/共82頁The pathway of light and the st

22、ages in magnification of a compound microscope第51頁/共82頁Effects of magnification第52頁/共82頁The parts of a laboratory microscope第53頁/共82頁Principle of Light Microscopy The magnifying power of the objectives alone usually ranges from 4to 100,and the power of the ocular（視覺的） alone ranges from 10to 20.The t

23、otal power of magnification of the final image formed by the combined lenses is a product of the separate powers of th two lens（透鏡）:第54頁/共82頁 Power of objective Usual power of ocularTotalmagnification=第55頁/共82頁Resolution（分辨力） Resolution is th capacity of an optical system to distinguish or separate

24、two adjacent objects or points from one another.第56頁/共82頁Resolving（分辨率）conditionresolvingNaked eyes0.2mmoptical microscope 0.2melectron microscope 0.2nm第57頁/共82頁.=Wavelength of light in nm（波長）/2*Numerical aperture(數(shù)值孔徑)Numerical aperture=n*sin a/2n:折射率第58頁/共82頁Effect of wavelength on resoltion第59頁/共

25、82頁Working of an oil immersion lens第60頁/共82頁PREPARING SPECIMENSFOR OPTICAL MICROSCOPES The condition of the specimen,either in a living or preserved(保存) state; The aims of the examiner,whether to observe overall structure,identify the micororganism,or see movment； The type of microsopy abailable,whe

26、ther is is bright-fiele(暗視野),phase-contrast（相差）,or fluorescence（熒光）.第61頁/共82頁Preparing specimens（標本）Fresh,Living PreparationsFixed（固定）,Stained（染色） SmearsSimple（簡單） Versus Differential（鑒別） Staining第62頁/共82頁Slide(載玻片)第63頁/共82頁Cover glass(蓋玻片)第64頁/共82頁第65頁/共82頁The making of smear（血涂片）第66頁/共82頁The flow

27、of optical microscope（顯微鏡觀察流程）Spreading a thin filmDryingFixationStainingObserving第67頁/共82頁Dark-Field Microscope(暗視野顯微鏡光路圖)第68頁/共82頁Negative Versue Positive Staining Positive stain:the dys actually sticks to the specimen and gives it color; Negative stain:For bright field microscopy, negative staini

28、ng is typically performed using a black ink fluid such as nigrosin.(苯胺黑) The specimen, such as a wet bacterial culture spread on a glass slide, is mixed with the negative stain and allowed to dry. When viewed with the microscope the bacterial cells, and perhaps their spores, appear light against the dark surrounding background 第69頁/共82頁Types of microbiological stains