LECTURES IN MICROBIOLOGY Microbial Nutrition and Growth LESSON

LECTURES IN MICROBIOLOGY Microbial Nutrition and Growth LESSON 5 Sofronio Agustin Professor

Lesson 5 Topics § Microbial Nutrition § Environmental Factors § Microbial Growth 2

Microbial Nutrition § Based on intake: (a) Macronutrients (CHONPS) (b) Micronutrients (trace elements) § Based on carbon content: (a) Organic nutrients- contain carbon (b) Inorganic nutrients- simple atom or molecule without carbon 3

Chemical Composition Bacteria are composed of different elements and molecules, with water (70%) and proteins (15%) being the most abundant. 4

Essential Nutrients §Carbon source §Energy Source §Growth Factors 5

Carbon Source § Autotrophs - obtain carbon from inorganic molecules like CO 2 § Heterotrophs - obtain carbon from organic matter from other life forms (e. g. sugar, proteins and lipids) 6

Energy Source §Photoautotrophs and photoheterotrophs obtain energy from sunlight §Chemoautotrophs derive electron energy from reduced inorganic compounds §Chemoheterotrophs obtain electron energy from hydrogen atoms of organic compounds 7

Nutritional Categories Summary of different nutritional categories of microbes based energy and carbon sources 8

Methanogens §Methanogens are chemoautotrophic microbes §Example: methane producing Archaea 9

Extracellular Digestion 10

Cell Membrane §Phospholipid bilayer with integral and peripheral proteins §“Fluid mosaic” model - phospholipids and proteins move laterally §Exhibits “selective permeability” 11

Membrane Transport § Passive: (a) Simple diffusion (b) Facilitated diffusion (c) Osmosis 1. Active: (a) Permease (b) Group translocation (c) Endocytosis 12

Simple Diffusion § Net movement of solute from area of high concentration to a low concentrated area § No energy is expended § Down the concentration gradient (like a river flowing downstream) 13

Diffusion A cube of sugar will diffuse from a concentrated area into a more dilute region, until an equilibrium is reached. 14

Facilitated Diffusion §Transport of polar molecules and ions across the membrane down their concentration gradients §No energy is expended (passive) §Carrier protein facilitates the binding and transport -Specificity -Saturation -Competition 15

Facilitated Diffusion: The Process 16

Osmosis § Diffusion of solvent (usually, water) through a permeable but selective membrane § Water tends to move toward higher solute concentrated areas 17

Tonicity Fate of cells in different osmotic conditions isotonic, hypotonic, and hypertonic solutions 18

Active Transport §Transport of molecules against its concentration gradient §Requires energy and transport protein (Ex. Permeases and protein pumps transport sugars, amino acids, organic acids, phosphates and metal ions) §Group translocation transports and modifies specific sugars 19

Endocytosis §Large substances are taken in by the cell but are not transported through the membrane. §Requires energy (active) §Common in eukaryotes - Phagocytosis - Pinocytosis 20

Active Transport Example of permease, group translocation and endocytosis 21

Cellular Transport : Summary 22

Environmental Factors § § § Temperature Gas p. H Osmotic pressure Other factors Microbial association 23

Temperature §Psychrophiles – (cold loving) 0 to 15 °C §Psychrotrophs - (food spoilage) grow between 20 to 30 °C §Mesophiles- (most human pathogens) 20 to 40 °C §Thermophiles- (heat loving) 45 to 80 °C §Themoduric - (contaminants of heated food) survive in short exposures to high temp §Hyperthermophiles - (Archaea) 24

Temperature Tolerance 25

Gas Requirements § Two gases that influence microbial growth: (1)Oxygen § Respiration - terminal electron acceptor § Oxidizing agent - toxic forms (2)Carbon dioxide 26

Oxygen Metabolites § Superoxide radical - O 2 § Singlet oxygen - O 2 with single electron in its valence shell § H 2 O 2 All are toxic byproducts of metabolism neutralized by enzymes SOD (superoxide dismutase), peroxidase and catalase. 27

Bacterial Types §Obligate aerobe §Facultative anaerobe §Obligate anaerobe 28

Obligate Aerobes §Require oxygen for metabolism §Possesses enzymes that can neutralize the toxic oxygen metabolites: SOD, peroxidase and catalase §Ex: Most fungi, protozoa, and bacteria like Bacillus sp. and Pseudomonas sp. 29

Obligate Anaerobes §Cannot use oxygen for metabolism §Do not possess SOD and catalase §The presence of oxygen is toxic to the cell §Ex: Clostridium sp. and Bacteroides sp. 30

Anaerobiosis Anaerobic culture techniques: (a) anaerobic chamber, (b) anaerobic jar 31

Facultative Anaerobes §Does not require oxygen for metabolism, but can grow in its presence §During minus oxygen states, anaerobic respiration or fermentation occurs §Possess superoxide dismutase and catalase §Ex. E. coli and S. aureus 32

Thioglycolate Broth Thioglycollate broth is used to demonstrate aerotolerance of bacteria. Aerobes, facultative anaerobes, and obligate anaerobes can be detected using this medium. 33

Other Gas Requirements § Microaerophiles - requires less than 10% of atmospheric O 2. Ex: Campylobacter jejuni § Capnophiles - requires increased CO 2 (5 -15%) tension for initial growth. Ex: S. pneumoniae 34

p. H §Most cells grow best between p. H 6 -8 §Acidophiles (up to p. H 0) - molds and yeast §Alkalinophiles (up p. H 10) ureadecomposing bacteria like Proteus sp. 35

Osmotic Pressure §Osmophiles - live in solutions with high solute concentration (e. g. sugar content in jams) §Halophiles - requires high salt concentrations and withstands hypertonic conditions Ex. Halobacterium sp. (Archaea) §Facultative halophiles - can survive high salt conditions but is not required for survival Ex. Staphylococcus aureus 36

Other Factors §Radiation- withstand UV, infrared rays §Barophiles – withstand high pressures §Spores and cysts- can survive dry habitats 37

Microbial Interactions Influence microorganisms have on other microbes: §Symbiotic relationship §Non-symbiotic relationship 38

Symbiotic Relationship Organisms that live together in close nutritional relationships Types: § Mutualism – both organism benefit § Commensalism – only one organisms benefits § Parasitism – typically host-microbe relationship 39

Commensalism § “Satellitism” as a form of commensalism § Staphylococcus aureus provides vitamins and amino acids to Haemophilus influenzae, which grows around colonies of S. aureus. 40

Non-Symbiotic Relationships § Organisms are free-living, and do not rely on each other for survival § Types: § Synergism – shared metabolism enhances growth of both microbes § Antagonism- competition between microorganisms 41

Microbe-Host Interactions §Can be commensal, parasitic, and synergistic §Ex. E. coli produce vitamin K for the host 42

Microbial Growth § Binary fission § Generation time § Growth curve § Enumeration of bacteria 43

Binary Fission §Parent cell enlarges and duplicates its DNA §Septum formation divides the cell into two separate chambers §Complete division results in two identical daughter cells 44

Steps in Binary Fission Rod-shaped bacteria undergoing binary fission 45

Growth Curve §Lag phase §Log phase §Stationary phase §Death phase 46

Phases of Bacterial Growth curve in a bacterial culture. 47

Enumeration of Bacteria § Direct Methods: (a) Microscopic (b) Viable plate count (c) Membrane filtration (d) Most probable number § Indirect Methods: (a) Turbidity (b) Metabolic assay (c) Dry weight determinations 48

Direct Microscopic Count §The direct cell method counts the total dead and live cells in a special microscopic slide containing a premeasured grid. §Petroff-Hausser counting chamber used in dairy industry. 49

Standard Plate Count Serially diluted samples are plated out and bacterial count expressed in CFU/ml. 50

Membrane Filtration Membrane filtration and coliform counts. 51

Turbidimetric measurements as indicators of bacterial growth. The greater the turbidity the larger the population density. 52

Coulter Counter §The Coulter Counter uses an electronic sensor to detect and count the number of cells. §Rapid automated counting method 53