1. Growth and replication of prokaryotes DR NAZIA KHAN

2. Requirements for Growth  Most of the dry weight of microorganisms is organic matter containing the elements carbon, hydrogen, nitrogen, oxygen, phosphorus, and sulfur. In addition, inorganic ions such as potassium, sodium, iron, magnesium, calcium, and chloride are required to facilitate enzymatic catalysis and to maintain chemical gradients across the cell membrane.  There must be a source of energy to establish the proton motive force and to allow macromolecular synthesis.  Microorganisms vary widely in their nutritional demands and their sources of metabolic energy.  Water(75-90% of the total body weight): source of carbon, nitrogen and some organic salts It participates in the metabolic reactions and also forms an integral part of the protoplasm

3. Bacteria can be classified nutritionally based on their energy requirement and their ability to form essential metabolites 1.Autotrophs:that can synthesize all their organic compounds,they are able to utilize atmospheric carbon dioxide and nitrogen. Not of much medical importance. Reduce carbon dioxide at the expense of water 2.Heterotrophs: unable to synthesize their own metabolites. require organic carbon for growth, and the organic carbon must be in a form that can be assimilated 3.Phototrophs: derive energy from light 4.Chemotrophs: derive energy from chemical reactions

4. Categories of requirement for microbial growth 1. Chemical requirments: includes sources of carbon, nitrogen, sulfur, phosphorus, trace elements, oxygen, and organic factors A.Major elements(macroelements or macronutrients) are the ones that make up cell constituents and over 95% of cell dry weight is made up of these. It includes carbon, oxygen, hydrogen, nitrogen, sulfur, phosphorus, potassium, magnesium, calcium, iron B.Trace elements(micronutrients): required in miunute quantities. They include cobalt, zinc, copper, molybdenum and manganese. They are part of enzymes Growth factors(bacterial vitamins):it is an organic compound which a cell must contain in order to grow, but it is unable to synthesis. They may be essential or accessory Example: vitamin B group, thiamine, riboflavin, nicotinic acid, pyridoxine,folic acid and vitamin b12

5. . 2. Physical factors influencing microbial growth: i.Temperature ii.Oxygen iii.Carbon dioxide iv.Moisture and drying v.pH vi.Light vii.Osmotic effect viii.Mechanical and sonic stresses

6. i.Temperature: the optimum temperature is 37°C. A.Mesophiles:grow best at 30–37 °C B.Psychrophiles: grow best at low temperatures (15–20 °C); C.Thermophilic:grow best at 50–60 °C D.Hyperthermophilic:grow at well above the temperature of boiling water, which exists under high pressure in the depths of the ocean.  Most organisms are mesophilic; 30 °C is optimal for many free-living forms, and the body temperature of the host is optimal for symbionts of warm-blooded animals

7. ii.Oxygen: A.Aerobes require oxygen for growth Obligate example: cholera vibrio Facultative: example: staphylococcus: most pathogenic bacteria are in this group Microaerophilic : grow best at low oxygen tension(-5%) example: campylobacter A.Anaerobes. Example: clostridium

8. iii.Carbon dioxide:All bacteria require some amount of carbon dioxide which they get from atmosphere or produce endogenously by cellular metabolism. Capnophilic bacteria: require higher levels(5-10%) of carbon dioxide example: Brucella abortus, pneumococcus, gonococcus iv.Moisture and drying: as water is essential,drying may lead to death of the bacteria v.pH: Most organisms (neutralophiles) grow best at a pH of 6.0–8.0, although some forms (acidophiles) have optima as low as pH 3.0 and others (alkaliphiles) have optima as high as pH 10.5.

9. iv.Light: darkness provides a favorable condition for growth and viability of bacteria. Exposure to light may influence pigment production. v.Osmotic effect: bacteria are more tolerant to osmotic variation because of the mechanical strength of the cell wall PLASMOLYSIS: when microorganisms with rigid cell walls are placed in a hypertonic solution, water leaves and plasma membrane shrinks away from the wall. This occurs more readily in gram negative bacteria than gram positive bacteria PLASMOPTYSIS: Sudden transfer of bacteria from concentrated solution to distilled water may also cause plasmoptysis due to excessive osmotic iimbibition of water leading to selling and rupture of the cell Mechanical and sonic stresses: they may be ruptured by vigorous shaking with glass beads.

10. Growth curve of prokaryotes: If a suitable liquid medium is inoculated with bacterium and incubated, its growth follows a definitive course The phases of the bacterial growth curve shown here are reflections of the events in a population of cells, not in individual cells. This type of culture is referred to as a batch culture. The typical growth curve may be discussed in terms of four phases 1.Lag phase:The lag phase represents a period during which the cells, depleted of metabolites and enzymes as the result of the unfavorable conditions that existed at the end of their previous culture history, adapt to their new environment express specific genes and synthesize components necessary for rapid growth 2.Exponential or logarithmic phase: binary fission occurs at a maximum rate, cells are dividing as rapidly as possible. This continues until one of two things happens: either one or more nutrients in the medium become exhausted, or toxic metabolic products accumulate and inhibit growth

11. 3. Stationary phase: Eventually, the exhaustion of nutrients or the accumulation of toxic products causes growth to cease completely. There is a slow loss of cells through death, which is just balanced by the formation of new cells through growth and division. When this occurs, the total cell count slowly increases although the viable count stays constant 4. Decline phase: After a period of time in the stationary phase, which varies with the organism and with the culture conditions, the death rate increases until it reaches a steady level. In most cases the rate of cell death is much slower than that of exponential growth. There is net decrease in the number of cells

12. Association of growth curve and cell changes The various stages of the growth curve are associated with morphological and physiological alterations of the cells. 1.Lag phase: bacteria have maximum cell size towards the end of the log phase 2.Log phase: cells are smaller and stain uniformly in the log phase. 3.Stationary phase: cells are frequently gram variable and show irregular staining due to the presence of intracellular storage granules. Sporulation occurs at this stage and also many bacteria produce secondary metabolic products such as exotoxins and antibiotics. 4.Decline phase: involution forms are common

13. Methods of replication in prokaryotes: Bacterial Reproduction: Asexual  Most bacteria reproduce by binary fission. During binary fission, the single DNA molecule replicates and both copies attach to the cell membrane. binary fission  The cell membrane begins to grow between the two DNA molecules.  Once the bacterium just about doubles its original size, the cell membrane begins to pinch inward.  A cell wall then forms between the two DNA molecules dividing the original cell into two

14. Self assessment 1.Mesophiles are the organisms which grow best at A.30–37 °C B.low temperatures (15–20 °C); C.50–60 °C D.above 100 °C 2.Phototrophs are the ones which a.derive energy from chemical reactions b.derive energy from light c.Cant synthesis their own metabolites d.None 3.Prokaryotes divide by a.Sexual reproduction b.Asexual reproduction c.Binary fission d.Both b and c 1.Chemotrophs: