SMALL-SCALE PROCESSING OF MICROBIAL PESTICIDES

Table of Contents

by
Professor V. Taborsky
University of Agriculture
Prague, Czechoslovakia

FAO AGRICULTURAL SERVICES BULLETIN No. 96

Food and Agriculture Organization of the United Nations Rome 1992

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Contents

FOREWORD

ACKNOWLEDGEMENTS

CHAPTER 1. GENERAL INTRODUCTION TO PROCESS OF MICROBIAL PESTICIDE
1.1 The Importance of Microbial Pesticides
1.2 Production and Commercialization of Pathogens
      1.2.1 Steps Leading to Commercialization
      1.2.2 Process Development and Production
               1.2.2.1 Organism Storage
               1.2.2.2 Fermentation Method
1.3 Theoretical Background of Industrial Processing in Biotechnology
      1.3.1 Laboratory Processing
               1.3.1.1 Cleanliness and Safety
               1.3.1.2 Preparation of Dilution
               1.3.1.3 Pure Culture Techniques Tube transfers
               1.3.1.4 Preparation and Testing of Culture Media
               1.3.1.5 Sterilization
                           1.3.1.5.1 Steam Sterilization
                           1.3.1.5.2 Flaming
                           1.3.1.5.3 Hot Air
      1.3.2 Preservation of Strains
      1.3.3 Elaboration of a Processing
               1.3.3.1 Laboratory Scale
               1.3.3.2 Small Scale Production
                           1.3.3.2.1 Problems of Contamination of Microbial Processes
                           1.3.3.2.2 Sterility of Microbial Process
                           1.3.3.2.3 Sensitivity of Microbial Processes to and Protection Against Contamination
     1.3.4 Pilot-Plant-Fermenters

CHAPTER 2. STANDARDIZATION OF PRODUCTS
2.1 Define the Objectives 
               2.1.1 What to Produce? 
               2.1.2 Which Propagule?
               2.1.3 Growth Factors
               2.1.4 Improving Mycoinsecticides
               2.1.5 Quality Assessment in Laboratory-Reared Insects 
2.2 Microbial Insecticide Based on Bacillus thuringiensis
               2.2.1 Morphology of Bacillus thuringiensis
                            2.2.1.1 Insecticidal Toxins Produced by Bacillus thuringiensis
                            2.2.1.2 The Role of Bacteriophages
                            2.2.1.3 Mode of Action
                            2.2.1.4 Taxonomy
                            2.2.1.5 Isolation of b-Exotoxin
                            2.2.1.6 Recovery of Bacillus thuringiensis
                            2.2.1.7 Bioassay Procedures
2.3 Microbial Insecticide Based on Metarhizium anisopliae
                  2.3.1 Mode of Action
                  2.3.2 Strain Improvement
                  2.3.3 Small-Scale Production
2.4 Microbial Insecticide Based on Beauveria bassiana
                  2.4.1 Small Scale Production
2.5 Microbial Insecticides Based on Verticillium lecanii
                  2.5.1 Effect of Humidity
                  2.5.2 Virulence and Strain Specific
                  2.5.3 Longevity Spores
                  2.5.4 Control of Aphids
                  2.5.5 Mode of Action Penetration of the Host Integument
                  2.5.6 Small-Scale Production
                  2.5.7 Formulated Product
                  2.5.8 Standardization
                  2.5.9 Registration

CHAPTER 3. DESCRIPTION OF THE PROCESSING
3.1 Small Scale Processing of Submerged Fermentation
                 3.1.1 Growth Kinetics in Submerged Culture
                 3.1.2 Factors Governing Sporulation
                 3.1.3 Mass Production
                 3.1.4 Production of Bacillus thuringiensis
                            3.1.4.1 Storage of Production Strain
                            3.1.4.2 Cultivation Media
                            3.1.4.3 Laboratory Scale Cultivation
                            3.1.4.4 Cultivation in Fermentation Tanks
                            3.1.4.5 Harvesting of Active Biomass
                            3.1.4.6 Final Adjustment
3.2 Small Scale Processing of Semi-Solid Fermentation
                 3.2.1 Production
                            3.2.1.1 Laboratory Cultivation of Inocula (seed culture)
                            3.2.1.2 Small Scale Processing
3.3 Small Scale Processing of Microbial Insecticides
                3.3.1 Processing of Microbial Insecticide on Solid Media
                            3.3.1.1 Introduction
                            3.3.1.2 Preparing Seed Inoculum
                            3.3.1.3 Processing by Cultivation
                            3.3.1.4 Formulation of Product

CHAPTER 4. PRODUCT HARVESTING AND FORMULATION OF MICROBIAL INSECTICIDES
4.1 Product Harvesting
4.2 Formulation
                4.2.1 Definitions
                4.2.2 Additives
4.3 Oil Suspension Formulation
4.4 Dusts or Wettable Powder
4.5 Suspension Concentrates (sc)
                4.5.1 Processing
                4.5.2 Function of the Surfactants
                4.5.3 Wetting
                4.5.4 Milling Aid
                            4.5.4.1 Stabilization
                            4.5.4.2 Milling Conditions
4.6 Suggested Evaluation Technique of Flowables
                4.6.1 Suspendibility
                4.6.2 Storage Stability
                4.6.3 Viscosity
                4.6.4 Bloom
                4.6.5 Biological Activity
4.7 Evaluation of Separation Process "Recovery"

CHAPTER 5. LABELLING AND PACKAGING

CHAPTER 6. TYPE OF EQUIPMENT IN THE LABORATORY
6.1 Introduction
6.2 Pilot Plant
               6.2.1 General Description
               6.2.2 Floor Areas
                           6.2.2.1 Wet Floor
                           6.2.2.2 Dry Floor
6.3 Laboratory Equipment
               6.3.1 Shake Flasks and Bottles
               6.3.2 Shaker Tables
               6.3.3 Shake Flask Volume
               6.3.4 Fermentation Glassware
               6.3.5 Stirred Fermenter
                           6.3.5.1 Automation and Performance
                           6.3.5.2 Agitation and Aeration
               6.3.6 Air
               6.3.7 Steam
               6.3.8 Water
               6.3.9 Electricity
               6.3.10 Gas
6.4 Fermenters
6.5 Ancillary Equipment
               6.5.1 Autoclaves
                           6.5.1.1 Bench-top or Portable Autoclaves
                           6.5.1.2 Standard Autoclaves
               6.5.2 Incubators, Shakers
               6.5.3 Ovens
               6.5.4 Pumps
                          6.5.4.1 Minipumps
                          6.5.4.2 Larger Pumps
6.6 Medium Reservoirs
6.7 Cooling System
6.8 "O" - rings

ANNEXES
ANNEX 1 - PROCESSING DEPARTMENTS
ANNEX 2 - CHARACTERISTIC OF FERMENTERS
ANNEX 3 - MAMPOWER
ANNEX 4 - REARING HELIOTHIS ZEA / HELIOTHIS VIRESCENS
ANNEX 5 - REARING OF TRICHOPLUSIA NI
ANNEX 6 - REARING OF SPODOPTERA EXIGUA