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CHAPTER Ⅰ GENERAL BACTERIOLOGICAL APPARATUS AND PROCEDURES1

Quality of Glass1

Hard Glass2

Soft Glass2

Cleaning Glassware2

Culture Tubes3

Petri Dishes4

Dilution Bottles and＂Water Blanks＂6

Fermentation Tubes7

Baskets and Culture Tube Racks10

Slides and Cover Glasses10

Needles and Loops13

Pipettes14

Plugging Tubes，Flasks，etc15

Incubators16

Miscellaneous Apparatus17

Sterilization and Disinfection17

Dry Heat Methods18

Incineration19

Flaming19

Hot Air Sterilization19

Moist Heat20

High Pressure Steam22

Boiling23

Free Flowing Steam23

Sterilization by Filtration24

Preparation and Storage of Sterile Apparatus26

CHAPTER Ⅱ THE MICROSCOPE AND ITS USE28

General Information28

General Construction of the Microscope28

Mechanical Parts29

The Body with its Draw Tube29

The Stage30

The Pillar31

The Coarse Adjustment31

Care of the Coarse Adjustment31

The Fine Adjustment31

The Substage Diaphragm32

Optical Parts33

Magnification33

Optical Glass33

The Eyepieces33

Which Eye to Use33

The Objectives34

Cover Glasses36

To Attach the Objective36

The Oil-immersion Objective37

Care of Objectives and Eye Pieces37

The Condenser38

Purpose of the Condenser38

Centering of Illumination39

To Focus Condenser39

The Mirror41

Use of the Microscope41

Finding an Object42

Micrometry—Measurement of Microorganisms46

CHAPTER Ⅲ CULTURE MEDIA50

General Remarks50

Standard Methods50

Characteristics of a Good Medium51

Constituents of Media51

Water51

Agar52

Classification of Sugars53

Gelatin54

Peptones54

Use of Indicators in Media55

Classification of Bacteriological Culture Media56

Liquid Media56

Beef Extract Broth56

Beef Infusion Broth57

Carbohydrate Broths57

Nitrate Broth58

Stiritz＇s Malt Medium for Yeasts58

Plain Milk58

Litmus Milk58

Brom-cresol-purple Milk58

Whey Broth59

Synthetic Media59

Liquefiable Solid Media59

Beef Extract Agar59

Wort Agar for Yeasts61

Lactic Acid Dextrose Agar for Yeasts61

Gorodkowa＇s Agar Medium for Yeasts62

Maneval＇s Modification of Gorodkowa＇s Agar Medium62

Beef Extract Gelatin62

Plain Gelatin63

Carbohydrate Gelatin63

Potato Slants63

Carrot Slants63

Dehydrated Media64

Filling Tubes，etc64

Reaction of Media and Cultures64

Indicators65

How is Reaction Expressed66

Titration Methods66

Measurement of Hydrogen Ion Concentration67

The Electrometric Method67

The Colorimetric Method69

Action of Buffers，＂Buffer Index＂70

Method for Adjusting the Reaction of Culture Media，etc71

Sterilization of Culture Media73

Storage of Culture Media74

References on Media and their Preparation74

CHAPTER Ⅳ MICROSCOPIC EXAMINATION OF BACTERIA75

Examination in Unstained Preparations75

Hanging Drop Preparations75

Lindner＇s Adhesion Culture76

India Ink Preparations78

Dark Field Illumination Examination78

Examination in Stained Preparation80

Stains and their Properties80

Mordants82

Theory of Staining82

Preparation of Apparatus83

Staining Solutions85

Preparation of Film for Staining85

General Microbiological Stains86

Aqueous Alcoholic Stains86

The Gram Stain86

Hucker＇s Modification of the Gram Stain89

Burke＇s Modification of the Gram Stain89

Reaction of Some Common Bacteria to the Gram Stain91

The Spore Stain93

Neisser＇s Method for Staining Spores93

Chromic Acid Method for Staining Spores93

May＇s Spore Staining Procedure93

Capsule Stains94

Muir＇s Method94

Welch＇s Method94

Huntoon＇s Method94

Hiss＇s Method95

The Gins Method of Demonstrating Capsules95

Acid Fast Staining97

Flagella Stains97

Loeffler＇s Flagella Stain98

Plimmer and Paine＇s Modification of the Casares-Gil Flagella Staining Procedure99

Van Ermengem＇s Flagella Stain100

Staining of Granules100

Preservation of Stained Preparations101

Differential Counting of Living and Dead Bacteria102

Bibliography102

CHAPTER Ⅴ ISOLATION OF BACTERIA103

Pure Cultures103

Dilutions103

Qualiative Method of Dilution104

Dilution in Liquefiable Solid Media105

Quantitative Method of Dilution105

Preparation of Petri Dish Cultures106

Method of Isolation of Pure Cultures107

Inoculation of Liquid Media109

Preparation of Stab Cultures109

Preparation of Shake Cultures110

Enumeration of Bacteria110

Collection of Samples111

Standard Agar Plate111

Streak Plates112

Disposal of Cultures113

Anaerobic Methods114

Fermentation Tube114

Buchner＇s Method114

Wright＇s Method115

Replacement of Oxygen by Hydrogen115

Dick＇s Method115

Methods Using Ordinary Petri Dishes116

Torrey＇s Method116

Krumwiede and Pratt＇s Method116

Frost＇s Method for the Study of Bacteria117

CHAPTER Ⅵ PROCEDURES FOR STUDYING THE CHARACTERISTICS OF BACTERIA121

Invigoration of Cultures121

Sketches122

Study of Morphological Characteristics122

Vegetative Cells122

Abnormal Forms123

Motility124

Endospores125

Capsules127

Study of Physiological Characteristics128

Cultural Characteristics128

Oxygen Relations128

Staining Reactions131

Nutrient Broth Cultures132

Agar and Gelatin Colonies134

Gelatin Stab135

Growth on Potato and the Hydrolysis of Starch136

Temperature Relations139

Thermal Death Point139

Action of Bacteria on Milk140

Litmus Milk，Purple Milk，etc141

Production of Hydrogen Sulfide143

Nitrate Reduction144

Detection of Indol Formation145

Salkowski Test145

Ehrlich＇s Test145

Gore Modification of Ehrlich＇s Test145

The Oxalic Acid Test for Indol146

Fermentation Reactions146

Detection of Gas Formation146

Acid Formation147

Pathogenicity to Animals148

The Index Number148

Agglutination Technic148

The Microscopic Agglutination Reaction149

The Macroscopic Agglutination Reaction150

Study of an Unknown Organism150

CHAPTER Ⅶ STUDY OF YEASTS152

Outline of Procedures for Studying Yeasts152

Detection of Ascospore Formation by Yeasts153

Hansen＇s Gypsum Block Method153

Maneval＇s Method153

Staining of the Nucleus in Yeasts154

Wager＇s Method154

Demonstration of Glycogen in Yeasts155

Demonstration of Fat in Yeasts155

Exercise：Study of Compressed Yeast155

Exercise：Preparation of Pure Culture of Yeast by Lindner＇s Method156

Exercise：Demonstration of Budding of Yeasts156

Exercise：Determination of the Fermenting Power of Yeasts156

Exercise：Determination of the Fermenting Activity of Yeasts157

Exercise：The Effect of Freezing on Yeasts158

CHAPTER Ⅷ STUDY OF MOLDS159

Outline of Observations to be Made on Molds159

Himmel＇s Method for Studying Mold Mycelium159

Exercise：Isolation of Oidium Lactis160

Exercise：Isolation of Rhizopus nigricans160

Exercise：Observations on Germination of Mold Spores160

Exercise：Effect of Heat on Mold Spores160

Exercise：The Effect of Freezing upon Mold Spores161

CHAPTER Ⅸ PLANS FOR WORKING163

Outlines for Studying Microorganisms in Classes163

CHAPTER Ⅹ SPECIAL EXERCISES196

Exercise：Characteristics of Unknown Organisms196

Exercise：Bacteriological Examination of Milk196

Exercise：Effect of Drying on Bacteria197

Exercise：Effect of Sunlight on Bacteria197

Exercise：Action of Disinfectants on Bacteria198

Exercise：Bacteriological Examination of Water199

Exercise：Determination of the Bacterial Content of Rain Water and Snow200

Exercise：Determination of the Number and Types of Bacteria in Ice200

Exercise：The Isolation and Study of Thermophilic Bacteria201

Exercise：Molds on Bread，Fruits，etc201

Exercise：Biological Test for Arsenic201

Exercise：Bacteriological Examination of Dish Water202

Exercise：To Determine the Types of Bacteria in Milk202

Exercise：Study of the Influence of Freezing on Bacteria202

Exercise：Isolation of Rhizobium leguminosarum(Bacillus radicicola)203

Exercise：Study of the Types of Curd Formed in Milk203

Exercise：Study of Bacteria in the Mouth203

Exercise：Effect of Dyes on Bacteria204

Exercise：Study of Bacteria in Air，Dust，etc204

Exercise：Bacteria on the Hands205

Exercise：Isolation of Bacteria from Boils or Pimples205

Exercise：Bacteria on Coins and the Oligodynamic Action of Metals205

Exercise：Examination of Candy206

APPENDIX211

Preparation of Reagents211

Normal Sodium Hydroxide211

Normal Hydrochloric Acid211

Preparation of Staining Solutions212

Methylene Blue—Saturated Aqueous Solution212

Methylene Blue—Saturated Alcoholic Solution212

Loeffler＇s Methylene Blue212

Gentian Violet—Saturated Aqueous Solution212

Gentian Violet—Stabilized(Stovall and Nichols，1916)213

Gentian Violet—Stabilized(Kilduffe，1909)213

Gentian Violet—Saturated Alcoholic Solution213

Carbol Gentian Violet213

Aniline Water Gentian Violet213

Fuchsin Basic—Saturated Aqueous Solution214

Fuchsin Basic—Saturated Alcoholic Solution214

Carbol Fuchsin214

Eosin—Aqueous Solution214

Eosin—Alcoholic Solution214

Safranine—Aqueous Solution214

Neutral Red—Aqueous Solution214

Bismarck Brown—Aqueous Solution215

Haematoxylin215

Lugol＇s Iodine Solution(Gram＇s)215

Core Test for Indol，Reagents215

Metric Equivalents216

Azolitmin Solution217

Andrade＇s Indicator217

Solutions for La Motte Comparator Sets217

Solutions for La Motte Hydrogen-ion Testing Sets218

Preparation of Indicator Solutions219

Indicators for the Determination of Hydrogen-ion Concentration219

Comparison of Thermometer Scales222

Index231