Unit 4: Lesson 1-Requirements for Growth

Aerotolerant Anaerobes

Aerotolerant Anaerobes- can't use O2 for growth but tolerate it well. -many ferment carbs to lactic acid -as lactic acid accumulates- it inhibits growth of aerobic competitors- establishes a favorable ecological niche for lactic acid producers -eg- lactobacilli- used in production of many acidic fermented foods (pickles and cheese) -make no use of oxygen in the air -can tolerate oxygen because they possess SOD or something that neutralizes that toxic forms of O2 previously discussed

10. NAME IT A prokaryotic cell hitched a ride to Earth on a space shuttle from some unknown planet. The organism is a psychrophile, an obligate halophile, and an obligate aerobe. Based on the characteristics of the microbe, describe the planet.

Cold, salty, aerobic

7.Nitrogen and phosphorus added to beaches following an oil spill encourage the growth of natural oil-degrading bacteria. Explain why the bacteria do not grow if nitrogen and phosphorus are not added.

Petroleum can meet the carbon and energy requirements for an oil-degrading bacterium; however, nitrogen and phosphate are usually not available in large quantities. Nitrogen and phosphate are essential for making proteins, phospholipids, nucleic acids, and ATP.

Physical Requirements

Temperature, pH, Osmotic Pressure

Summary

The Requirements for Growth (pp. 154-160) 1. The growth of a population is an increase in the number of cells. 2. The requirements for microbial growth are both physical and chemical. Physical Requirements (pp. 154-158) 3. On the basis of preferred temperature ranges, microbes are classified as psychrophiles (cold-loving), mesophiles (moderate-temperature-loving), and thermophiles (heat-loving). 4. The minimum growth temperature is the lowest temperature at which a species will grow, the optimum growth temperature is the temperature at which it grows best, and the maximum growth temperature is the highest temperature at which growth is possible. 5. Most bacteria grow best at a pH value between 6.5 and 7.5. 6. In a hypertonic solution, most microbes undergo plasmolysis; halophiles can tolerate high salt concentrations. Chemical Requirements (pp. 158-160) 7. All organisms require a carbon source; chemoheterotrophs use an organic molecule, and autotrophs typically use carbon dioxide. 8. Nitrogen is needed for protein and nucleic acid synthesis. Nitrogen can be obtained from the decomposition of proteins or from NH4+ or NO3−; a few bacteria are capable of nitrogen (N2) fixation. 9. On the basis of oxygen requirements, organisms are classified as obligate aerobes, facultative anaerobes, obligate anaerobes, aerotolerant anaerobes, and microaerophiles. 10. Aerobes, facultative anaerobes, and aerotolerant anaerobes must have the enzymes superoxide dismutase open parenthesis 2 O sub 2 to the times below − power plus 2 H to the plus power rightwards arrow O sub 2 plus H sub 2 O sub 2 close parenthesis and either catalase open parenthesis 2 H sub 2 O sub 2 rightwards arrow 2 H sub 2 O plus O sub 2 close parenthesis or peroxidase open parenthesis H sub 2 O sub 2 plus 2 H to the plus power rightwards arrow 2 plus H sub 2 O close parenthesis . 11. Other chemicals required for microbial growth include sulfur, phosphorus, trace elements, and, for some microorganisms, organic growth factors. Biofilms (pp. 160-161) 1. Microbes adhere to surfaces and accumulate as biofilms on solid surfaces in contact with water. 2. Biofilms form on teeth, contact lenses, and catheters. 3. Microbes in biofilms are more resistant to antibiotics than are free-swimming microbes.

Lesson Notes

While some of the chemicals that microorganisms need for growth are used in minute quantities (trace elements), others are essential sources of continued sustenance for microorganisms. The section "Recycling Vital Elements" (p. 15) describes two major chemical cycles used by some microorganisms, namely the nitrogen cycle and the carbon cycle. Use of these molecules, as this section shows, is essential for the maintenance and growth of the microorganisms.

How organisms can be harmed by oxygen- requires brief discussion of the toxic forms of oxygen.

• 1. Singlet oxygen (1O2−) is normal molecular oxygen (O2) that has been boosted into a higher-energy state and is extremely reactive. 2. Superoxide radicals (O2-), or superoxide anions, are formed in small amounts during the normal respiration of organisms that use oxygen as a final electron acceptor, forming water. In the presence of oxygen, obligate anaerobes also appear to form some superoxide radicals, which are so toxic to cellular components that all organisms attempting to grow in atmospheric oxygen must produce an enzyme, superoxide dismutase (SOD), to neutralize them. Their toxicity is caused by their great instability, which leads them to steal an electron from a neighboring molecule, which in turn becomes a radical and steals an electron, and so on. Aerobic bacteria, facultative anaerobes growing aerobically, and aerotolerant anaerobes (discussed shortly) produce SOD, with which they convert the superoxide radical into molecular oxygen (O2) and hydrogen peroxide (H2O2): 3. 3. The hydrogen peroxide produced in this reaction contains the peroxide anion O22− and is also toxic. InChapter 7 (page 199) we will encounter it as the active principle in the antimicrobial agents hydrogen peroxide and benzoyl peroxide. Because the hydrogen peroxide produced during normal aerobic respiration is toxic, microbes have developed enzymes to neutralize it. The most familiar of these is catalase, which converts it into water and oxygen: 2 H2O2 → 2 H2O + O2 Catalase is easily detected by its action on hydrogen peroxide. When a drop of hydrogen peroxide is added to a colony of bacterial cells producing catalase, oxygen bubbles are released. Anyone who has put hydrogen peroxide on a wound will recognize that cells in human tissue also contain catalase. The other enzyme that breaks down hydrogen peroxide is peroxidase, which differs from catalase in that its reaction does not produce oxygen: H2O2 + 2 H+ → 2 H2O. Another important form of reactive oxygen, ozone (O3), is also discussed on page 199. 4. 4. The hydroxyl radical (OH·) is another intermediate form of oxygen and probably the most reactive. It is formed in the cellular cytoplasm by ionizing radiation. Most aerobic respiration produces traces of hydroxyl radicals, but they are transient. -these toxic forms of oxygen- essential component of one of the body's most important defenses against pathogens- phagocytosis -in phagolysosome of the phagocytic cell- ingested pathogens are killed by exposure to singlet oxygen, superoxide radicals, peroxide anions of hydrogen peroxide, and hydroxyl radicals and other oxidative compounds -obligate anaerobes- usually produce neither superoxide dismutase nor catalase -because aerobic conditions lead to an accumulation of superoxide radicals in their cytoplasm- obligate anaerobes are extremely sensitive to oxygen

The effect of oxygen on the growth of various types of bacteria

* important

typical growth rates of different types of microorganisms in response to temperature

*see Figure 6.1 in notes****

pH

-acidity or alkalinity of a solution -most bacteria grow best in a narrow pH range between 6.5-7.5 -few grow in acidic pH below 4 -hence, why sauerkraut, pickles, and cheeses are preserved from spoilage by acids produced by bacterial fermentation -acidophiles-bacteria that are tolerant of acidity -1 type of chemoautotrophic bacteria- found in drainage waterfrom coal mines- oxidizes sulfur to form sulfuric acid- can survive at pH 1 -molds & yeasts- grow over a greater pH range than bacteria, but optimum pH of molds and yeasts is generally below that of bacteria (usually pH 5 or 6) -Alkalinity-also inhibits microbial growth- but is rarely used to preserve foods -bacteria cultured in lab- often produce acids that interfere w/ their own growth -chemical buffers are included in growth medium -peptones & amino acids- act as buffers -many media also contain phosphate salts- have an advantage of exhibiting their buffering effect in the pH growth range of most bacteria. Are also nontoxic- provide phosphorus (an essential nutrient)

Microaerophils

-aerobic -require oxygen -only grow in oxygen conc. lower than those in air. - In a test tube of solid nutrient medium, they grow only at a depth where small amounts of oxygen have diffused into the medium; they do not grow near the oxygen-rich surface or below the narrow zone of adequate oxygen. -limited tolerance -due to their sensitivity to superoxide radicals and peroxides- which they produce in lethal conc. under O2 rich conditions.

Nitrogen, Sulfur, and Phosphorus

-also needed to synthesize cellular material -Nitrogen & some sulfur- needed for protein synthesis -Nitrogen & some phosphorus- synthesis of DNA & RNA, synthesis of ATP -Nitrogen- makes up 14% of dry weight of bacterial cell - Sulfur & Phosphorus together- constitute another 4% -use nitrogen to form amino group of amino acids of proteins - Many bacteria meet this requirement by decomposing protein-containing material and reincorporating the amino acids into newly synthesized proteins and other nitrogen-containing compounds. -other bacteria- use N from ammonium ions (NH4+)- already in reduced form & are usually found in organic cellular material -other bacteria- derive nitrogen from nitrates (compounds that dissociate to give the nitrate ion, NO3- in olution. -some bacteria (photosynthesizing cyanobacteria)- use gaseous nitrogen (N2) directly from atmospherecalled nitrogen fixation -used by both the plant and the bacterium (symbiosis) -Sulfur- used to synthesize sulfur-containing amino acids & vitamins such as thiamine and biotin -important natural source of sulfur: sulfate ion (SO42-), hydrogen sulfide, and sulfur-containing amino acids. -Phosphorus- essential for synthesis of nucleic acids & phospholipids of cell membranes. -also found in energy bonds of ATP -source of phosphorus: Phosphate ion (PO43-) -Potassium, magnesium, and calcium- elements that microorganisms require, often as cofactors for enzymes

Temperature

-certain bacteria can grow at extremes of temps that might kill eukaryotic organisms -microorganisms- classified into 3 groups on preferred range of temp: -Psychrophils- love COLD -Mesophiles- love MODERATE TEMPERATURE -Thermophiles- love HOT -most bacteria- grow within a limited range of temps (max and min growth temps are only about 30 degrees apart) -grow poorly at high and low temp extremes within their range- -grow at minimum, optimum, and max temperatures -Minimum Growth Temp- lowest temp in which species will grow -Optimum Growth Temp- temp where species grow best -Maximum Growth Temp- highest temp at which growth is possible -higher than optimum growth temp- growth rates drop rapidly b/c high temps can inactivate necessary enzymatic systems of the cell.

Biofilms

-communities of microorganisms -reside in a matrix made up primarily of polysaccharides- also containing DNA and proteins- called SLIME -biofilm can also be considered a HYDROGEL -Hydrogel-a complex polymer containing many times its dry weight in water -Quorum Sensing-cell-to-cell communications -allows bacteria to coordinate their activity and group together into communities- provide benefits -biofilms aren't just slime layers, but biological systems -bacteria are organized into a coordinated, functional community -usually attached to a surface (rock, human tooth), or a mucous membrane -might be of single species or of a diverse group of microorganisms -Might take other, more varied forms- floc that forms in certain types of sewage treatment (eg) -within biofilm community- bacteria are able to share nutrients, are sheltered from harmful factors of env (desiccation, antibiotics, and body's immune system) -close proximity of microorganisms within a biofilm may also have advantage of facilitating transfer of genetic info by conjugation -biofilm begins- when a free swimming (planktonic) bacterium attaches to a surface -f these bacteria grew in a uniformly thick monolayer, they would become overcrowded, nutrients would not be available in lower depths, and toxic wastes could accumulate -avoid this by forming pillar-like structures w/channels between them - water can carry incoming nutrients and outgoing wastes- primitive circulatory system -microbes in biofilms- work cooperatively to carry out complex tasks -essential elements in proper functioning of sewage treatments -can also be a problem in pipes and tubing- accumulations impeded circulation -biofilms- important factor in human health -microbes in biofilms- 1000 times more resistance to microbicides -70% of human bacterial infections involve biofilms -most nosocomial infections (infections acquired in health care facilities) are related to biofilms on medical catheters -can include biofilms formed by fungi such as Candida- encountered by infections related to use of contact lenses, dental caries, and infections by pseudomonad bacteria -preventing biofilmformation- incorporate antimicrobials into surfaces on which biofilms might form. -chemical signals that allow quorum sensing- essential to biofilm formation- research is trying to determine the makeup of these chemical signals to perhaps block them -another approach- discovery of lactoferrin- abundant in human secretions- can inhibit biofilm formation -Lactoferrin- binds iron (esp among pseudomonads responsible for cystic fibrosis biofilms) -lack of iron- inhibits the surface motility essential for aggregation of the bacteria into biofilms -most lab methods in microbiology- use organisms being cultured in their planktonic mode -predict that there will be an increasing focus on how microorganisms actually live in relation with one another - this will be considered in industrial and medical research.

Organic Growth Factors

-essential organic compounds an organism is unable to synthesize - organic growth factors -must be directly obtained from env. -group of OGF for humans- vitamins -most vitamins function as coenzymes- the organic cofactors required by certain enzymes in order to function -many bacteria can synthesize their own vitamins- don't depend on outside sources -some bacteria lack the enzyme needed for synthesis of certain vitamins- for them, those vitamins are OGF. -other OGF required for some bacteria- Amino acids, Purines, Pyrimidines.

Halophiles

-extreme halophiles- have adapted to high salt concentrations- they actually require them for growth. -may be termed obligate halophiles -eg. Organisms from saline waters as the dead sea -facultative halophiles- more common- do not require high salt conc. but are able to grow at salt conc. up to 2%, a conc. that inhibits growth of other organisms -a few facultative halophiles can tolerate even 15% salt -most microbes- must be grown in a medium that is nearly all water -eg. Conc of agar (a complex polysaccharide isolated from marine algae)- used to solidify microbial growth- about 1.5% salt -if higher conc. are used- increased osmotic pressure can inhibit growth -if osmotic pressure is unusually low- environment is hypotonic (eg as in distilled water) -water tends to enter the cell rather than leave it -microbes w/ a weak cell wall may be lysed by such treatment.

Chemical Requirements: Carbon

-important for microbial growth (besides water) -structural backbone of living matter -needed for all the organic compounds that make up a living cell -Chemoheterotrophs- get most of their carbon from the source of their energy- organic materials such as: PROTEINS, CARBS, and LIPIDS -Chemoautotrophs, and Photoautotrophs- derive carbon from CO2 -Carbon makes up 50% of dry weight of bacterial cell

Trace Elements

-iron, copper, molybdenum, zinc-trace elements -most essential for functions of certain enzymes- usually as cofactors -usually assumed to be naturally present in tap water and other components of media -most distilled water- contain adequate amounts -tap water- sometimes specified to ensure that trace minerals will be present in culture media

Microbial Growth

-microbes that are growing are increasing in number -accumulate into colonies (groups of cells large enough to be seen without a microscope - become hundreds of thousands of cells or populations of billions of cells -bacteria can survive & grow slowly in nutrient-poor environments by forming biofilms. -2 main requirements for microbial growth: physical and chemical -Physical requirements for microbial growth: Temperature, pH, and osmotic pressure -Chemical requirements for microbial growth: sources of carbon, nitrogen, sulfur, phosphorus, oxygen, trace elements, and organic growth factors

Osmotic Pressure

-microorganisms obtain almost all nutrients in solution from the surrounding water -require water for growth -composition is 80-90% water -high osmotic pressures- have the effect of removing necessary water from a cell -when amicrobial cell is in a solution whose conc. of solutes is higher than that in the cell (environment is hypertonic to the cell), the cellular water passes out through the plasma membrane to the high solute concentration. -this osmotic loss of water causes plasmolysis - shrinkage of the cell's cytoplasm -importance: the growth of the cell is inhibited as the plasma membrane pulls away from the cell wall -thus- addition of salts (or other solutes) to a solution resulting increase in osmotic pressure can be used to preserve foods. -eg: salted fish, honey, and sweetened condensed milk -the high salt or sugar concentrations draw water out of any microbial cells that are present- preventing their growth. -effects of osmotic pressure- roughly related to the number of dissolved molecules and ions in a volume of solution

Mesophiles

-optimum growth temp (25-40 degrees) -most common type of microbe -optimum temp for many pathogenic bacteria- about 37 degrees C -include most of the common spoilage and disease organisms

Thermophiles

-optimum growth temp of 50-60 degrees C -can be reached in sunlit soil, and thermal waters (hotsprings) -cannot grow at temps below 45 degrees C -endospores formed by themophilic bacteria- unusually heat resistant & may survive the usual heat treatment given canned goods. -elevated storage temps- may cause surviving endospores to germinate and grow- spoiling food- these thermophilic bacteria are not considered a public health problem -Thermophiles- important in organic compost piles -Archaea- optimum growth temp of 80 degrees or higher -called hyperthermophiles or extreme thermophiles -live in hot springs associated w/ volcanic activity -sulfur is usually important in their metabolic activity -bacterial growth and replication at high temps (known record) is about 121 degrees C near deep-sea hydrothermal vents -immense pressure in ocean depths prevents water from boiling

Psychrophiles

-psychrophiles, mesophiles, or thermophiles aren't rigidly defined. Psychrophiles -2 fairly distinct groups capable of growing at 0 degrees celcius. -1 group: composed of psychrophiles in the strictest sense: -can grow at 0degrees C -optimum growth temp of 15 degrees C -so sensitive to high temps- will not grow in a reasonably warm room -found in oceans' depths, or certain polar regions -seldom cause problems in food preservation -other group: can grow at 0degrees -has higher optimum temps (20-30degrees C)- cant grow above 40 degrees -much more common than psychrophiles -likely to be encountered in low-temp food spoilage- grow well at refrigerator temps -Psychrotrophs- favored by food microbiologists, for this type of spoilage microorganisms -Psycotrophs-an organism capable of growth between 0-30 degrees celcius. -Refrigeration- common method of preserving household food supplies -based on principle: microbial reproductive rates decrease at low tmeps -microbes usually survive even subfreezing temps (might become dormant), but gradually decline in number. -Psycotrophs- do not grow well at low temps- except in comparison w/ other organisms; -given time- they are able to slowly degrade food -temperature inside a properly set fridge will greatly slow the growth of most spoilage organisms and will entirely prevent the growth of all but a few pathogenic bacteria

Oxygen

-usually think of molecular oxygen (O2) as necessity to life- -actually in a sense is a poisonous gas -many current forms of life have metabolic systems that require oxygen for aerobic respiration -H atoms that have been stripped from organic compounds, combine w/ oxygen to form water -this process yields a great deal of energy- while neutralizing a potentially toxic gas -Microbes that use molecular oxygen (aerobes) extract more energy from nutrients than microbes that don't use oxygen (anaerobes) -Obligate Aerobes- organisms that require oxygen to live -Obligate Aerobes- at a disadvantage b/c oxygen is poorly soluble in the water of their env. -many of the aerobic bacteria have developed, or retained, the ability to continue growing in the absence of oxygen. -Facultative anaerobes -Facultative Anaerobes- can use oxygen when its present but are able to continue growth by using FERMENTATION or ANAEROBIC RESPIRATION when O2 isnt avail. -efficiency in producing energy DECREASES in absence of O2 -eg. Escherichia coli (found in human instestinal tract) -eg. Many yeasts -many microbes are able to substitute other electron acceptors- such as nitrate ions, for O2 (in anaerobic respiration), which is something humans cant do. -Obligate Anaerobes- bacteria that are unable to use O2 for energy-yielding reactions -most are harmed by it -genus Clostridium (contains species that cause tetanus and botulism) -do not use O2 atoms present in cellular materials- atoms are obtained from water.

Objectives

1. list the possible physical and chemical requirements for growth. 2. define psychrophile, mesophile, and thermophile. 3. define optimum growth temperature. 4. compare the physical properties of acidophiles and halophiles. 5. explain the importance of carbon, nitrogen, sulphur, and phosphorus as requirements for microbial growth. 6. compare the oxygen requirements of obligate aerobes, facultative anaerobes, obligate anaerobes, and microaerophiles. 7. demonstrate an understanding of the basis of toxicity of oxygen intermediates for microorganisms.