Introdução Lab

Question 1

Sterilization methods

Answer 1

Sterilization= The killing or removal of all viable organisms within a growth medium

-Radiation
Ionizing radiation and non-ionizing radiatio; (U.V.) is used for sterilization in the medical field and food industry

• Heat sterilization: the most widely used method of controlling microbial growth –> High temperatures denature macromolecules
• -> Dry Heat sterilization: 180ºC, 2h
• ->Calor Húmido: autoclave (sealed device: steam under pressure, allows T of water > 100ºC): 121ºC, 1 atm, 15min
• Filtration: avoids the use of heat on sensitive liquids and gases; membrane filters (0.45 μm; 0.22 μm)
• Flaming

Question 2

Pasteurization

Answer 2

process of using precisely controlled heat to reduce the microbial load in heat-sensitive liquids

Does not kill all organisms–> different from sterilization!

Some bacteria produce resistant cells = endospores
–> Can survive heat that would rapidly kill vegetative cells!

Question 3

Culture media

Answer 3

Nutrient solutions used to grow microbes in the lab

2 broad classes:

• Defined media: precise chemical composition is known
• Complex media: composed of digests of chemically undefined substances (e.g., yeast and meat extracts)

Nutrients
--> Supply of monomers (or precursors of) required by cells
for growth
• Macronutrients
--> Nutrients required in large amounts
• Micronutrients
--> Nutrients required in trace amounts

Question 4

Types of Media

Answer 4

• Simple media: contains only essential nutrients (Glucose- fonte de C/energia; K2HPO4/KH2PO4- tp, fonte de K e P; (NH4)2SO4, MgSO4.7H20; CaCl2; Co, Mn,Zn…H20)
• Complex media: contains all the elements most bacteria need for growth + is non-selective –> general cultivation and maintenance of bacteria kept in lab culture collections (Glucose; Extrato de levedura- fonte de fatores de crescimento; Peptona- fonte de AA, N, S; K2HPO4; H20)
• Selective medium: suppress the growth of some microorgs while allowing the growth of others (Halófilos Extremos - [sais] > 4 M)
• Differential medium: recognized on the basis of their colony colour. Various approaches include incorporation of dyes, metabolic substrates… –> bacteria that utilize them appear as differently coloured colonies.

Question 5

Growth-Dependent Diagnostic Methods

Answer 5

a) Use of Differential Medium to Assess Sugar Fermentation (precipita?/ turbidez)
b) Conventional Diagnostic Test for Enteric Bacteria in Triple Sugar Iron (TSI) Agar

Question 6

Measuring Microbial Growth

Answer 6

• Viable Cell Counting
• Measurements of Microbial Mass: Turbidimetric Methods
• Measurements of Total Cell Numbers: Microscopic Counts

Question 7

Antibiogram

Answer 7

Inoculate plate w/ liquid culture of test org in nutrient agar plate –> antibiotic discs are placed on surface –> incubate for 24-48h

Test organism shows susceptibility to some antibiotics, indicated by inhibition of bacterial growth around discs