Microbial growth and metabolism

Nutrition, Growth, and Metabolism1In 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 geneticsMicrobial growth requirementsWhat are the chemical growth factors required for isolation of microbes in vitro?

1-CarbonOrganic sourceGlucoseInorganic sourceCO2

2-Nitrogen Organic sourceProteinInorganic sourceAtmospheric nitrogen

Autotrophic microbes using inorganic carbon and nitrogenHeterotrophic microbes using organic carbon and nitrogen

Other chemical requirements3- Hydrogen, Oxygen, Phosphorus, Carbon, Nitrogen, Ions, and Sulfur

Physical growth factors1- pH and Buffer requirementsPathogenic bacteria grow best at neutral pH which is typically between pH (6.8 to 7.4)

Acidophilic bacterium is able to survive with acidic pH Example: Helicobacter pylori

On the other extreme, bacteria that prefer alkaline conditions are known as alkaliphilesExample: Vibrio cholerae

Fungi such as yeasts and molds; grow best at acidic conditions ( pH 5 )2- Salts concentration

A specific concentration of NaCl is required for microbial growth in vitroIt is equal to normal saline salts concentration (0.9% NaCl)Halophilic bacteria resist high salt concentration3- Temperature requirements

Mesophilesgrow at optimum of 37 Chuman body temperature Pathogens Opportunists Pyschrophileclose to freezing

Thermophileclose to boiling

4- Gaseous requirements and Humidity According to oxygen requirements, microbes can be divided into the following groups

1- Strict or Obligate AerobeO2 must be present to growExample: Mycobacterium sp.

2- Strict or Obligate AnaerobesOxygen must be absentKilled by oxygenExample: Clostridium sp. 3- Facultative anaerobesAdaptable organisms that use oxygen when present but can switch to anaerobic pathways in its absenceSurvive in the presence of O2Example: Escherichia coli

4- MicroaerophilicOnly use low concentrations of oxygen ( around 5%)Example : Helicobacter pylori

Grow in low oxygenKilled in high oxygen

The Bacterial Growth CurveBacterial 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

During stationary phase, the growth rate slows as a result of nutrient depletion and 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

Bacterial binary fission

Bacterial growth curveMicrobial MetabolismAccording to biochemical pathway used in energy production, bacterial metabolism can be categorized into three types:

1-Aerobic RespirationMolecular oxygen serves as the final electron acceptor38 ATP molecules will be produced by oxidation of one glucose moleculeUsed by obligatory aerobic bacteria for energy productionsuch as: Mycobacterium sp.

2-Anaerobic RespirationInorganic sulfate or nitrate act as the final electron acceptor38 ATP molecules will be produced by catabolism of one glucose moleculeUsed by obligatory anaerobic bacteria such as: Clostridium sp.

3-FermentationLactic acid ( produced by bacteria) or ethanol ( produced by yeast) serves as final electron acceptorOnly 2 ATP molecules will be produced by fermentation of one glucose moleculeUsed by facultative anaerobic bacteria such as : E. coli

Microbial GeneticsProkaryotic 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

Both bacteria chromosome and plasmid are called replicons

Genetics is the study of inheritance and variation

Genetic information encoded in DNA

Function of genetic material1- Replication of the genome2- Expression of DNA to mRNA then to protein

DefinitionsGenotypethe complete set of genetic determinants of an organism

Phenotypeexpression of specific genetic material

Bacterial DNA2 types of DNA in bacteria

ChromosomalExtra-chromosomal (plasmid)

PlasmidExtrachromosomal DNAFound in most species of bacteria.Govern their own replicationGenetic exchange, amplify genesTransfer by conjugationCode for resistance to antibiotics & toxins

Bacterial Structure

Bacterial StructureGene TransferTransfer 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

Transduction is the process by which DNA is transferred from one bacterium to another by a virus22Bacterial conjugationConjugation: 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

Conjugation24TransductionTransduction is the process by which DNA is transferred from one bacterium to another by a virus

It also refers to the process whereby foreign DNA is introduced into another cell via a viral vector

TransductionBacterial transformationA stable genetic change brought about by the uptake of naked DNA and competence refers to the state of being able to take up exogenous DNA from the environment

There are two forms of transformation and competence: natural and artificialThe Microbial Virulence factorsVirulence factors are external cellular structures, enzymes, and toxins that enhance microbial pathogenicity

In general, the most important virulence factors are:

1-Microbial capsuleMicrobe resist host acidic environment (stomach gastric acid)Microbe resist host proteolytic enzyme (Present in Saliva, and stomach)Microbe resist phagocytosis2-Fimbriae or PiliMicrobial adhesion to the host cell surfaceAdhesion could be also enhanced by receptor-antigen interaction

3-Microbial EnzymesCollagenase enzymeenhances microbial invasion; due to degradation of extracellular matrix componentsUreaseNeutralization of acidic pH ( urine, stomach) CoagulaseCatalase have different functions4-Bacterial ToxinsA-ExotoxinsWell known poisonous substances.Chemical natureProteins (two polypeptide components)Almost all are Heat-labile at 60 C

Intracellular toxin fraction could1-Inhibit cellular protein biosynthesis2-Cause ionic imbalance and loss of water3-Inhibit the release of neurotransmitters

B-EndotoxinsChemical natureLipopolysaccharide, the component of Grams negative bacterial outer membraneHeat-Stable at 100 C

5-The microbial HemolysinDegradation of RBCs, Hemoglobin and NADH will be released

6-The microbial Haemagglutinine and Coagulase enzymeAgglutination of RBCs; the microbe escapes Humoral immunity7-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

In the absence of -lactam antibiotics, the bacterial cell wall plays an important role in bacterial reproduction.Adding -lactam antibiotics to the cell medium while bacteria are dividing will cause them to shed their cell walls and fail to divide, forming large, fragile spheroplasts.