第四章 微生物的营养和培养基 47

1、 Chapter 4Nutrition and medium of microbes(微生物的营养和培养基微生物的营养和培养基)Nutrition： the most basic physiological function for organisms to intake the energy and material that is necessary for living and reproduction from the outside environment .(营养是指生物体从外部环境中摄取对其生命活动必需的能量和物质,以满足正常生长和繁殖需要的一种最基本的生理功能.)Nutrien

2、t: it is referred to as the nutritional material including the unconventional material formray radiation in microbiology.(营养物则是指具有营养功能的物质,在微生物学中,它还非常规物质形式的光辐射能在内.) Nutriton and nutrient Section A six nutritional elements of microbes(微生物的六类营养要素微生物的六类营养要素) The chemical composition of the microorganism

3、 cells and their metabolite determines what nutrient they need.As bacteria are a very disparate group of microbes that can exist in a wide range of environments, and can be split into a number of different nutritional groups with very different prerequisites for growth, the ingredients in the medium

4、 will depend on the individual species. The common requirements of all bacteria are water, a source of energy, carbon, nitrogen, essential inorganic ions such as phosphate, sulfate, sodium, calcium, magnesium, potassium and iron and a number of trace elements (Zn, Mn, Mo, Se, Co, Cu, Ni, W). For het

5、erotrophs, energy and carbon can be derived from the same molecule. The composition of the microbe cell carbon, hydrogen, oxygen , nitrogen and essential inorganic ions such as phosphate, sulfate, sodium, calcium, magnesium, potassium and iron and a number of trace elements (Zn, Mn, Mo, Se, Co, Cu,

6、Ni, W): water,carbohydrates, protein, nucleic acid, lipoid, vitamine, antibiotic , inorganic ion.Elemental levelCompound level1.Carbon source2.Nitrogen source3.Energy source4.Growth factor 5.Mineral salts6.Water Level of Nutritional elementsWhat does a microbe need for growth?Carbon source are defin

7、ed as all the nutrients that can satisfy the need for carbon during the process of microbial growth. It is also called macronutrients(大量营养物大量营养物) in that it is in the most greatest need only secondary to water. The spectrum of carbon sources (碳源谱碳源谱)are extremely wide.The spectrum of carbon sources

8、can be divided into two types: organic carbon and inorganic carbon. Those who have to depend on the organic carbon source are heterotrophs, the majority of microorganisms are of this type.And those who use inorganic carbon source as main form are autotrophs. Only a few of microbes are of this type.C

9、arbon sourceHeterotrophs and autotrophs For all the heterotrophs, the carbon source is also energy source at the same time, therefore this kind of carbon resources is called difunctional nutrient(双功能营养物双功能营养物).Nitrogen source provide the microorganism with the element of nitrogen for its living and

10、reproduction. The spectrum of nitrogen source is also quite wide and have two types: organic nitrogen and inorganic nitrogen.In the culture medium the commonly used organic nitrogen are beef extract, yeast extract and peptone(牛肉膏牛肉膏, 酵母膏和蛋酵母膏和蛋白胨白胨).Nitrogen sourceAmino acid autotrophs(氨基酸自养型生物氨基酸自养

11、型生物) is a group of organism who dont need amino acid as their nitrogen source and can self-synthesize all the amino acids they need using simple nitrogen source as material.Otherwise, amino acid heterotrophs (氨基酸异养型生物氨基酸异养型生物) intake ready amino acids as their nitrogen resources . All the animals an

12、d a number of heterotrophs are amino acids heterotrophs.Amino acid autotrophs and amino acid heterotrophs Energy source are those nutrients that provide the initial energy or ray radiation energy for its living. The spectrum of energy source can be divided into two types: chemical material(organic o

13、r inorganic) and irradiation energy. Chemotrophs use chemical materials and phototrophs use irradiation energy.Energy sourceMonofunctional nutrient)ray irradiation energy （ 单功能营养物单功能营养物 energy source）Bifunctional nutrient)deoxidized（还原态（还原态 的）的）inorganic NH4 (双功能营养物双功能营养物（energy source and nitrogen

14、source） Trifunctional nutrient)amino acids (三功能营养物三功能营养物 source of carbon， nitrogen and energy）A certain nutrient might has two or three nutritional functions，that is ,it can be used as the source of carbon， nitrogen or energy，while some only have one function .Growth factor Growth factor is a kind

15、of organic material cant be self -synthesized by auxotrophs(营养缺陷型微生物营养缺陷型微生物) with simple carbon source and nitrogen source, while is necessary for microorganisms to regulate the normal metabolism.Although growth factor is an important nutrient element, it is different from the carbon source , energ

16、y source and nitrogen source in that not any certain organism need the growth factor provided by the environment. Bacteria that can synthesize all they require from these basic ingredients are called prototrophs（原养型微生物原养型微生物） and most microorganisms that survive in the outside environment can do thi

17、s.PrototrophsMicrobes that have become adapted to life in a situation rich with nutrients such as the human body may require other growth factors to be provided, such as vitamins, amino acids or nitrogenous bases. These organisms are called auxotrophs（营养缺陷型）. Leuconostoc mesenterdides, for example,

18、requires over 40 additional growth factors. Bacterial growth media are therefore designed to suit the organism of interest and may be either defined or complex media.auxotrophsMicrobes can be divided into three types on the basis of situation of the need for growth factor: Auxoheterotrophs(生长因子异养性微生

19、物生长因子异养性微生物) Auxoautotrophs (生长因子自养性微生物生长因子自养性微生物) Growth factor over synthesized microbes(生长因子过量生长因子过量 合成的微生物合成的微生物)Normally, lactic bacteria needs vitamines as growth factor, many microbes and their auxotroph mutant strains need nitrogenous base.Some natural material contains rich growth factors t

20、herefore can act as growth factor source such as yeast extract, cornsteep liquor(玉米浆玉米浆),liver infusion(肝浸液肝浸液), malt extract (麦麦芽汁芽汁) etal.Some microbes has the ability of secrete certain vitamines etal. in large amount during the metabolism, thus can be used as vitamine-producing bacterial strains

21、.Mineral saltsMineral salts or inorganic salts(无机盐或矿质元素无机盐或矿质元素) supply the important elements except the carbon source and nitrogen source. macroelements大量元素大量元素 10-310-4 mol/LMineral salts microelements微量元素微量元素 10-610-8 mol/L As far as the macroelement is concerned, when a medium is prepared ,abov

22、e all we usually select K2HPO4 and MgSO4, for they can provide four elements that the microbes need in the greatest amount at the same time. Normally enough trace elements are present in the water.Section B Nutritional type of microbe Photoautotrophs (光能自养微生物光能自养微生物) Photoheterotrophs (光能异养微生物光能异养微生

23、物) Chemoautotrophs (化能自养微生物化能自养微生物) Chemoheterotrophs (化能异养微生物化能异养微生物) saprophytism(腐生腐生)dead（saprophyte）Chemoheterotrophs parasitism (寄生寄生) living（parasite）寄生）寄生 菌菌 Obligatory parasitism Parasitism Facultative parasitism Facultative saprophytismSection C How the nutrients enter into the cell（营养物质进入

24、细胞的方式）（营养物质进入细胞的方式） Dont through the carrier protein of membrane ： Simple diffusionStyles dont consunme energy ：facilitated diffusion pass through the carrier protein active transport（untouched） consume energy group translocati（changed） Simple diffusionout of the membranemembranecytoplasmConformatio

25、n changesconformation comebackFacilitated diffusionActive transportConformation changes by consuming energyCome back to the original conformationEnzyme AEnzyme BGroup translocationclassification criteriumNutritional types1. energy sources 2 . H donator 3. Carbon sources4. amino-synthesis ability5. G

26、rowth factor6. Ways of intaking 7. Living or dead organics Section D Medium（培养基）（培养基） Medium is an artificial mixture containing all the six appropriated essential nutritional elements (requirements ) for the growth and reproduction or the production of metabolites of a certain microbe .(培养基是指由人工配制的

27、适合微生物生长繁殖或产生代谢产物用的混合营养料. 任何培养基都具备微生物生长所需要的六大营养要素 且期间的比例是合适的)1.Principles and methods for the medium selection and designing (1) Four principles A . Definite aim 目的明确 B. Harmony (appropriate) nutrient 营养协调 C. proper physical and chemical condition 理化适宜 pH, osmotic pressure, redox potential (氧化还原电势) D

28、. economical 经济节约1.Principles and methods for the medium selection and designing Four methodsA ecology simulation 生态模拟生态模拟B literature referring 参阅文献参阅文献C self-designing 精心设计精心设计D comparative trying 实验比较实验比较 2. Medium classification2.1 On the basis of the knowledge of its ingredients: (按对培养基成分的了解分类按

29、对培养基成分的了解分类)A complex media (undefined media)B synthetic media (chemical defined media)C semi-synthetic media (semi-defined media ) 2. Medium classification2.2 On the basis of physiological condition (按培养基外观的物理状态分类按培养基外观的物理状态分类)A liquid mediaB solid mediaC semi-solid mediaD dehydrated culture media

30、(dehydrated commercial media or pre-fabricated dried culture media) 脱水商品培养基或预制干燥培养基脱水商品培养基或预制干燥培养基 solid media : solidified media (gelling agent: 12%agar, gelatin, alginate) irreversible solidified media (silica gel as gelling agent or serum media ) natural solid media Membrane filter (滤膜滤膜) 2. Medi

31、um classification2.3 On the basis of the function : (按培养基对微生物的功能分类按培养基对微生物的功能分类)A selected media (选择培养基选择培养基) enriched selected media (加富性选择培养基加富性选择培养基) inhibited selected media (抑制性选择培养基抑制性选择培养基)B differential media (鉴别培养基鉴别培养基) eg. Eosin Methylene Blue (EMB) Microbes are grown (cultured) in the la

32、boratory in either liquid or solid medium. Bacterial growth media should contain all the essential requirements for growth: an energy, carbon and nitrogen source as well as the essential ions, phosphate, sulfate, sodium, calcium, magnesium, potassium and iron, and various trace elements. Prototrophs

33、 require no additional ingredients in the medium, while auxotrophs require additional growth factors such as amino acids, vitamins and nitrogenous bases. The media used may be denfined (synthetic) media(限定，合成培养基限定，合成培养基) where all the ingredients are known or complex media（天然培养基）（天然培养基） containing a

34、 mixture of undefined nutrients. Agar is used to solidify a liquid medium. Selective or differential media are used to detect the presence of particular groups of microbes.Key Notesgrowth media Each bacterial species has an optimum temperature, oxygen concentration, pH and water activity for growth

35、and will survive in a range of conditions around these optima.Key NotesEnvironmental conditions for growth in the laboratory bacteria are normally grown (cultured) in either liquid medium, in flasks, bottles or large culture vessels called fermenters, or solid medium, in Petri dishes, which are roun

36、d, normally plastic, sterile dishes. Introduction of microbes into or on to these media is called inoculation. The nature of the medium depends on the microorganisms natural environment and on the reason why it is being grown. Normally, to isolate large numbers of microbes or their products, liquid

37、culture is used but solid medium is used for the isolation of individual bacteria and for storage. Liquid medium and solid medium Liquid medium (or broth) can be converted into solid medium by the addition of agar (1-2%) which is isolated from seaweed. It is a particularly useful polysaccharide poly

38、mer because, once it is melted by boiling, it does not harden until it is cooled to 40-42C which allows the addition of temperature-sensitive ingredients, such as proteins, before it sets. Having set, it will not melt again until it has been heated to 80-90C. Agar is also not normally degraded by ba

39、cteria. Petri dishes containing a nutrient agar are often referred to as agar plates.How broth becomes solid media？agar A defined（限定） or synthetic medium contains known ingredients which are usually the basic requirements for growth of a particular microbe. These are normally simple (minimal) media

40、that contain the minimal requirements for growth. Examples of defined media for the photosynthetic Cyanobacteria (note the lack of a carbon source) and heterotrophic E. coli are shown in Table .Defined mediumA complex medium contains undefined ingredients such as proteolytic digests of meat (peptone

41、s) and meat and yeast extracts which provide enough ingredients (amino acids, vitamins, sugars and bases) to sustain the growth of a wide range of microbes. Complex media such as nutrient broth (NB) and tryptic soya broth (TSB豆胨肉汤豆胨肉汤) are routinely used for cultivation of bacteria in the laboratory

42、 and are particularly useful for growing bacteria whose growth requirements have not been defined. Blood is frequently used as an additive to media, for the isolation of human pathogens, as it provides many of the essential nutrients for the growth of fastidious human pathogens such as Streptococcus.Complex medium1. Peptones2. meat and yeast extracts 3. nutrient broth (NB) 4. tryptic soya broth (TSB）5. bloodThe following substances are normally added