Microbial growth and metabolism:

1. Microbial growth and metabolism: In Medical Microbiology, microbial cultivation is required for the following purposes: In Medicine: 1-Diagnosis of most infectious diseases. 2-Selection of drug of choice ( antibiotics ) for treatment of infection. In other fields: 3-Preparation of Vaccine. 4-Research tool in molecular genetics.

2. Microbial growth requirements Chemical growth factors What are the chemical growth factors required for isolation of microbes in vitro? 1-Carbon: Organic source: Glucose Inorganic source: CO2 2-Nitrogen: Organic source: Protein Inorganic source: atmospheric nitrogen Autotrophic microbes using inorganic carbon and nitrogen. Heterotrophic microbes using organic carbon and nitrogen. Other chemical requirements 3- Hydrogen, Oxygen, Phosphorus , and Sulfur.

3. Physical growth factors: 1- pH and Buffer requirements: Pathogenic bacteria grow best at neutral pH which is typically between pH (6.8 to 7.4). -Acidophilic bacteriumis able to survive with acidic pH. Example:Helicobacter pylori -On the other extreme, bacteria that prefer alkaline conditions are known as alkaliphiles. Example: Vibrio cholerae. Fungisuch as yeasts and molds; grow best at acidic conditions ( pH 5 ).

4. a 2- Salts concentration: A specific concentration of NaCl is required for microbial growth in vitro. It is equal to normal saline salts concentration (0.9% NaCl). Halophilic bacteria resist high salt concentration. 3- Temperature requirements: Mesophiles:Grow at optimum of 37 ᵒC. Normal flora and most pathogens grow at or near human body temp. of 37 ᵒC .

5. 4- Gaseous requirements and Humidity : According to oxygen requirements, microbes can be divided into the following groups: 1- Strict ,Obligate Aerobe: O2 must be present. Example: Mycobacterium. 2-Strict or obligate Anaerobes: Molecular oxygen must be absent. Example:Clostridium. 3- Facultative anaerobes: Adaptable organisms that use oxygen when present but can switch to anaerobic pathways in its absence. Example: Escherichia coli. 4- Microaerophilic: Only use low concentrations of oxygen ( around 5%). Example : Helicobacter pylori.

6. The Bacterial Growth Curve: Bacterial growth is the division of one bacterium into two daughter cells in a process called binary fission. Bacterial growth curve represents the relationship between microbial quantity and time of incubation. During lag phase, bacteria adapt themselves to growth conditions (number of dividing cell is zero). Exponential phase( Log phase): is a period characterized by Rapid cell doubling. Doubling time can be as short as twenty minutes or as long as several days.

7. a During stationary phase, the growth rate slows as a result of nutrient depletionand accumulation of toxic products. This phase is a constant value as the rate of bacterial growth is equal to the rate of bacterial death. At death phase, bacteria run out of nutrients and die.

8. Microbial Metabolism: According to biochemical pathway used in energy production, bacterial metabolism can be categorized into three types: 1-Aerobic Respiration: -Molecular oxygen serves as the final electron acceptor. - 38 ATP molecules will be produced by oxidation of one glucose molecule. - Used by obligatory aerobic bacteria for energy production; such as: Mycobacterium.

9. n 2-Anaerobic Respiration: -Inorganic sulfate or nitrate act as the final electron acceptor. -38 ATP molecules will be produced by catabolism of one glucose molecule. -Used by obligatory anaerobic bacteria such as: Clostridium. 3-Fermentation: -Lactic acid ( produced by bacteria) or ethanol ( produced by yeast) serves as final electron acceptor. -Only 2 ATP molecules will be produced by fermentation of one glucose molecule. -Used by facultative anaerobic bacteria such as : E. coli.

10. Microbial Genetics: Prokaryotic Genome: -Most prokaryotic genes are carried on the bacterial chromosome, a single circle of DNA. -Many bacteria contain additional genes on plasmids. -Plasmid is an extra-circular supercoiled DNA that carry some important genes such as the antibiotics resistance genes. Bothbacteria chromosome and plasmid are called replicons.

11. Gene Transfer: -Transfer of DNA among prokaryotes is widespread between different strains of same bacterial species. Mechanisms of Gene Transfer: 1-Conjugation. 2-Transduction(is a phage-mediated genetic transfer). 3- Transformation. Conjugation: is a mechanism of gene transfer by which plasmids will be transferred from one bacterial cell to another by a mean of Sex pili. Hospital-dwelling bacteria resist antibiotics due to conjugation.

12. Conjugation: n

13. The Microbial Virulence factors: Virulence factors are external cellular structures, enzymes, and toxins that enhance microbial pathogenicity. In general, the most important virulence factors are: 1-Microbial capsule: Microbe resist host acidic environment (stomach gastric acid). Microbe resist host proteolytic enzyme, (Present in Saliva, and stomach). Microbe resist phagocytosis.

14. N 2-Fimbriae or Pili: -Microbial adhesion to the host cell surface. -Adhesion could be also enhanced by receptor-antigen interaction. 3-Microbial Enzymes: Collagenase enzyme enhances microbial invasion; due to degradation of extracellular matrix components. Urease: Neutralization of acidic pH ( urine, stomach). Coagulase, Catalase have different functions.

15. n 4-Bacterial Toxins: A-Exotoxins: - Well known poisonous substances. - Chemical nature: Proteins (two polypeptide components). - Almost all are Heat-labile at 60 ˚C. - Intracellular toxin fraction could: 1-inhibit cellular protein biosynthesis. 2-Cause ionic imbalance and loss of water. 3-inhibit the release of neurotransmitters.

16. n B-Endotoxins: -Chemical nature: Lipopolysaccharide, the component of Gram’s negative bacterial outer membrane. -Heat-Stable at 100 ˚C. 5-The microbial Hemolysin: -Degradation of RBCs, Hemoglobin and NADH will be released. 6-The microbial Haemagglutinine and Coagulase enzyme: -Agglutination of RBCs; the microbe escapes Humoral immunity.

17. n 7-The microbial Beta-Lactamases: -Some microbes have ability to resist antibiotics due to production of Beta-Lactamase enzymes. Some strains of Staphylococci can hydrolyze the Beta- Lactam ring shown.