Measurement of Bacterial of Growth
Measurement of Bacterial of Growth
Measurement of Bacterial of Growth
A convenient method is to determine turbidity by photoelectric colorimeter or spectrophotometer.
The cell number can be counted as total cell number as well as viable count. Viable Count Viable number of bacteria can be counted by inoculating the suspension onto solid growth medium and counting the number of colonies. Since each colony is the end product of one viable bacterium, their count gives the number of viable bacteria in the suspension.
Total number of bacteria can be ascertained in specially designed chambers such as Coulter counter.
Two methods are known for genetic variation in bacteria: mutation and gene transfer.
Mutation : Any change in the sequence of bases of DNA, irrespective of detectable changes in the cell phenotype. Mutations may be spontaneous or induced by various agents which are known as mutagens.
Spontaneous Mutations: Arise from enzymatic imperfections during DNA replications or with transient insertions of transposable elements.
Induced Mutations: Mutation by physical and chemical mutagens.
Physical mutagens ultraviolet rays and high-energy ionizing radiations. The primary effect of UV rays on DNA is the production of pyrmidine dimers whereas ionizing radiations cause single_stranded breaks the DNA molecules.
Chemical mutagens :Affecting nucleotide sequence
(i) Agents which cause error in base pairing (e.g. nitrous acid and alkylating agents).
(ii) Agents which cause errors in DNA replication (e.g. acridine dyes such as acridine orange and profiavine).
(iii) Base analogs which are incorporated into DNA and cause replication errors (e.g. 5-bromouracil)
Transformation: Uptake of naked DNA
Transduction : Infection by a nonlethal bacteriophage
Conjugation : Mating between cells in contact
Transformation: Gene transfer by soluble DNA is called as transformation. it requires that DNA be absorbed by the cell, gain entrance to the cytoplasm and undergo recombination with the host genome.
Artificial Transformation(transfection) :Some of the bacteria (such as Escherichia coli) resist transformation until they are subjected to some special treatment such as CaCl2 to make the bacterium more permeable to DNA. Such modified cells can also take up intact double stranded DNA extracted from viruses or in the shape of plasmids. Though the process is same as transformation, it is 9 as transfection because it results in infection by an abnormal route
Transduction :The type of gene transfer in which the DNA of one bacterial cell is introduced into another bacterial cell by viral infection is known as transduction. This introduces only a small fragment of DNA. Because the DNA is protected from damage by the surrounding phage coat, transduction is an easier to perform and more reproducible process than transduction. ,
Two types of transduction are known.
- Generalized transduction When a bacteriophage picks up fragments of host DNA at random and can transfer any genes
- Specialised transduction: phage DNA that has been integrated into the host chromosome is excised along with a few adjacent genes, which the phage can then transfer.
After entry into the host cell, the phage DNA gets incorporated into the host chromosome in such a way that the two genomes are linearly contiguous (lysogeny). The phage genome in this stage is known as prophage, The host cell acquires a significant new property as a consequence of lysogeny because it becomes immune to infection by homologous phage. This is hence called as lysogenic conversion and endow toxigenicity to Corynebacterium diphtheriae
Abortive Transduction :phage DNA fails to integrated into the host chromosome, the process is called as abortive transduction The phage DNA does not replicate and along with binary fission Of the host it goes into one of the daughter cells.
Conjugation :This is defined as the transfer of DNA directly from on bacterial. .cell to another by a mechanism that requires cell-to-cell contact.
The capacity to donate DNA depends upon the possession of the fertility (F) factor. The F pili also retard male-male union. Concomitant with effective male-female pair formation, the circular DNA bearing the F factor is converted to a linear form that is transferred to the female cell in a sequential manner. DNA replication occurs in the male cell and the newly synthesized, semiconserved DNA molecule remains in the male. This ensures postmating characters of the male.
Conjugation in Different Bacteria: Unusual form of plasmid transfer, called phase mediated conjugation has been reported to occur with some strains of Staphylococcus aureus.
Protoplast Fusion: Also called as genetic transfusion. Under osmotically buffered Conditions protoplast fusion takes place by joining of cell membrane and generation of cytoplasmic bridges through which genetic material can be exchanged.
Transposons: Transposons Tn are DNA sequences which are incapable of autonomous existence and which transpose blocks of genetic material back and forth between cell Chromosome and smaller replicons such as plasmids. insertion sequences (IS ) are another similar group of nucleotides which can move from one chromosome to another
Genetic material. IS and Tn are collectively also known as transposable elements or Jumping genes. These are now recognised to play an important role in bringing about vanous types of mutations.
NORMAL MICROBIAL FLORA
A. Properties. Normal microbial flora describes the population of microorganisms that usually reside in the body. The microbiological flora can be defined as either
1) Resident flora - A relatively fixed population that will repopulate if disturbed,
2) Transient flora - that are derived from the local environment. These microbes usually reside in the body without invasion and can
even prevent infection by more pathogenic organisms, a phenomenon known as bacterial interference.
The flora have commensal functions such as vitamin K synthesis. However, they may cause invasive disease in immunocompromised hosts or if displaced from their normal area.
B. Location. Microbial flora differ in composition depending on their anatomical locations and microenvironments. The distribution of normal microbial flora.
Mitochondrion – double MB structure responsible for cellular metabolism – powerhouse of the cell
Nucleus – controls synthetic activities and stores genetic information
Ribosome – site of mRNA attachment and amino acid assembly, protein synthesis
Endoplasmic reticulum – functions in intracellular transportation
Gogli apparatus/complex – composed of membranous sacs – involved in production of large CHO molecules & lysosomes
Lysosome – organelle contains hydrolytic enzymes necessary for intracellular digestion
Membrane bag containing digestive enzymes
Cellular food digestion – lysosome MB fuses w/ MB of food vacuole & squirts the enzymes inside. Digested food diffuses through the vacuole MB to enter the cell to be used for energy or growth. Lysosome MB keeps the cell iself from being digested
-Involved mostly in cells that like to phagocytose
-Involved in autolytic and digestive processes
-Formed when the Golgi complex packages up an especially large vesicle of digestive enzyme proteins
– vesicle that forms around a particle (bacterial or other) w/in the phagocyte that engulfed it
- Then separates from the cell membrane bag & fuses w/ lysozome to receive contents
- This coupling forms phagolysosomes in which digestion of the engulfed particle occurs
- Contain catalase
- Bounded by a single membrane bag
- Compartments specialized for specific metabolic pathways
- Similar in function to lysosomes, but are smaller & isolate metabolic reactions involving H2O2
- Two general families:
· Peroxisomes: transfer H2 to O2, producing H2O2 – generally not found in plants
· Glyoxysomes: common in fat-storing tissues of the germinating seeds of plants
¨ Contain enzymes that convert fats to sugar to make the energy stored in the oils of the seed available
– transitory, non-living metabolic byproducts found in the cytoplasm of the cell
- May appear as fat droplets, CHO accumulations, or engulfed foreign matter.
The cell cycle
1) Labile cells (GI tract, blood cells)
- Described as parenchymal cells that are normally found in the G0 phase that can be stimulated to enter the G1
- Undergo continuous replication, and the interval between two consecutive mitoses is designated as the cell cycle
- After division, the cells enter a gap phase (G1), in which they pursue their own specialized activities
• If they continue in the cycle, after passing the restriction point (R), they are committed to a new round of division
• The G1 phase is followed by a period of nuclear DNA synthesis (S) in which all chromosomes are replicated
• The S phase is followed by a short gap phase (G2) and then by mitosis
• After each cycle, one daughter cell will become committed to differentiation, and the other will continue cycling
2) Stable cells (Hepatocytes, Kidney)
- After mitosis, the cells take up their specialized functions (G0).
- They do not re-enter the cycle unless stimulated by the loss of other cells
3) Permanent cells (neurons)
- Become terminally differentiated after mitosis and cannot re-enter the cell cycle
- Which cells do not have the ability to differentiate -> Cardiac myocytes
Types of microscopy used in bacteriology
Phase contrast microscopy
Transmission electron microscopy
Scanning electron microscopy
Fluorescent microscopy in which ultraviolet rays are used to examine cells after treatment with fluorescent days.
Phase contrast microscope enhances the refractive index differences of the cell components. This microscopy can be used to reveal details of the internal structures as well as capsules, endospores and motility
Electron microscope The resolving power is more than 200 times that of light microscope.