Midterm 3

Symbiotic associations

**Ex: Rhizobium + root nodules of leguminous plants **Ex: Fungi + cyanobacterium = "lichen"

Hyperthermophiles

100C optimum growth temperature 68C-108C temp range

Thermophiles

> 65C optimum growth temperature 45C-80C temp range

So far we've covered two cell types of anabaena (vegetative and heterocysts). Whats the third type?

Akinete: thick-walled dormant cells for servival; granular cytoplasms

Volume of hemocytometer chamber (Grid)

Area is 1mm^2 Depth is 0.1mm Volume = 1mm^2 * 0.1 mm = 0.1 mm^3 = 0.0001 cm^3 = 0.0001 ml

Which plates would you use to calculate the concentration of a culture that was diluted as noted on the table? A)1 and 2 B) 3 and 4 C) 5 and 6 D) A and B E) All the plates

B. You take the average. Between 30-300 colonies per plate are considered countable colonies => Average (64+57)/2 = 60.5 = 61 (round .5 up to 1) => 61/0.1 ml = 610 cfu/ml = 6.1 x 10^2 in the diluted sample (10^-7) => CDF = 1/10^-7 => Concentration of original: 6.1 x 10^2 * (1/10-7) = 6.1 x 10^9 cfu/ml

Bacteriocidal vs Bacteriostatic

Both are antibiotics. Bacteriocidal kills bacteria by either interfering with formation of cell wall or cell contents Bacteriostatic stops bacteria from multiplying by interfering with: => protein production => DNA replication => or other aspects of cellular metabolism

Halotolerant

Can grow under saline conditions but do not require elevated concentrations of salt for growth.

- Free-living - Symbiotic associations

Environments: -soils, freshwater ponds, lakes and streams, coastal and open ocean marine environments

Free-living: Anabaena, cyanobacterium

Filamentous Cyanobacteria - blue/green bacteria

Lag phase

Growth of cell before division. Synthesis of enzymes, ATP, and increase in size.

Free-living diazotrophs

In free-living diazotrophs, the ammonia is further converted to glutamine through the glutamine synthetase pathway and incorporated into cellular material. **Ex: The cyanobacterium, Anabaena are capable of fixing nitrogen in soil or water environments

How can OD values be used to give you information on cell culture concentration?

In order to be able to use OD values as a measure of cell culture concentration, we have to first generate a standard curve. A standard curve correlates the concentration of a culture (cells/ml) to particular OD values. Then you can measure the OD value of the sample with unknown concentration and use the standard curve to extrapolate the concentration from the standard curve.

What can a spectrophotometer measure?

It can measure either transmitted light or absorbed light. We used the absorbance setting because Beer's law says that the amount of light absorbed by a medium is proportional to the concentration of the absorbing material. The absorbance is measured as the Optical Density (OD). So we get the absorbance value at OD600. Ex: OD600 = # <--this number is absorbance and it has no units.

What is the generation time/doubling time?

It is the time interval in which the population of cells doubles in each generation **Example: E. coli divides every 20 min. It's generation time is 20 min.

Leghemoglobin

Leghemoglobin is a protective compound that sequesters the surrounding oxygen, keeping oxygen levels low enough to allow nitrogenase to function but high enough to allow aerobic respiration.

The enzyme that catalyzes the reduction of dinitrogen to ammonia is?

Nitrogenase The nitrogenase reaction is an energy intensive process that requires: => 8 protons => 16 ATP

- Explain the various ways antibiotic resistance testing is standardized. Why is this important?

One must standardize such things as medium type and inoculum size. This is important because the size of the ZOI varies according to how easily the antibiotic diffuses through the agar, the type of medium used, and other factors. So we have to standardize the type of medium used and the inoculum size in order to relate the ZOI size to whether an isolate is resistant, sensitive, or intermediate in its response to a particular antibiotic.

How rhizobia protects its nitrogenase from oxygen?

Rhizobia is aerobic. What's the solution to the nitrogenase problem? The solution must keep oxygen levels sufficiently low to prevent the inactivation of nitrogenase and sufficienty high to support respiration. ***Solution: Leghemoglobin

What does OD value mean? What does it not mean?

The OD value gives you an indication of whether the density of the sample has changed or not. It does NOT tell you about cell numbers or concentration of the culture.

Our experiment: survival in extreme conditions

The control tube is TSB incubated at 30/37C (optimum temp of unk) => Media used for testing pH is selective. => Media used for testing salt is selective. => Media used for testing temp is NOT selective. The temp is a selective condition but the media is not.

The ecological importance of nitrogen fixation

There's an increased demand for fixed nitrogen due to increased need for food crops. => commerical synthetic nitrogen fertilizer is expensive to make and uses a lot of energy. Improper use can result in fertilizer running off of farms and polluting water bodies. => Current research is focused on increasing natural methods like increasing nitrogen fixation through genetic engineering, etc.

Osmoprotectant

These molecules accumulate in cells and balance the osmotic difference between cell's surroundings and cytosol

Halophiles

Thrives in high solute environments

Experiment for cell differentiation in anabaena

We enriched for nitrogen-fixing anabaena by inoculating: => BG11 minimal media: phosphate, NITRATE, no organic carbon +light => BG11-0 minimal media: phosphate, NO NITRATE, no organic carbon +light ***Note: because anabaena is photoautotroph, it will grow in media without organic carbon source (it makes its own complex organic carbons) using light as their energy source

How to make a standard curve

Y axis: OD values X axis: Concentration (cells/ml) Data points come from: => Take a sample of known concentration and measure its OD. => Measure OD values of different dilutions of that sample => Make various dilutions to get you several OD values between .1 and .8 Standard curves are specific to the particular culture and does not universally apply to other cultures. For example, the concentration of yeast cells at a given OD may be lower than the concentration of a bacterial culture at the same OD because of the difference in cell size between yeast and bacteria. **In this lab, we determined cell culture density of YEAST CELLS using spectrometery

Nitrogen fixation

converts atmospheric dinitrogen to ammonia and incorporates it into organic nitrogen **N2, the most abundent form of nitrogen, cannot be used by most organisms. Uptake of nitrogen is most efficient as ammonia or nitrate and most easily passed from one organism to another through the food chain as amines, nucleotides, and urea. Thus this process of nitrogen fixation is really important to the level of primary production.

What is a hemocytometer? How does it work? What are it's advantages/disadvantages?

It is a thick glass microscope slide that has a grid embedded into its surface. The size of the grid on the slide and the weight of the accompanying coverslip are extremely precise. The exact volume of the grid is known and this allows us to count the cells in a given volume to get the concentration (cells/ml) of the culture. **Benefits: Very precise for determining number of cells/ml and is also faster than the plating method **Disadvantages: Not as fast as spectrophotometer and it doesn't differentiate between dead and alive cells (bc we count both)

- Explain the Kirby Bauer method. How does it work?

Kirby-Bauer method is the standard recommended by the US food and drug admin. Things that are standardized in the Kirby Bauer method: => on Mueller-Hinton agar is used => chemical composition of agar is defined => pH always 7.2-7.4 => uniform thickness 4 mm => uniform moisture/hydration => thoroughly inoculated using a cotton swab from a broth culture having a turbidity that matches a defined standard => only discs containing certain contentrations of antibiotics are used => words used to describe the effectiveness of each antibiotic are rigidly defined (R,S,I)

How does a spectrophotometer work?

Light source sends light through lens, which converges light though a monochromator (prism) and the wavelength selector selects the wavelength it is set for. (We set it to 600nm). Then the monochromatic light passes through the sample solution and the fraction of light that is able to pass through is measured by a photometer.

In our experiment we screened for antibiotic producers among bacteria by examining the effect of those bacteria on the growth of 4 different target bacterial strains.

Plated four different 4X4 grid containing agar plate with one of the following bacteria strains: 1) Pseudomonas 2) E. coli 3) Bacillus 4) M. luteus Tapped a tooth pick in each square for each of the 15 antibiotic producers and unknown (16 total) Then we measured zone of inhibition (ZOI) in mm from the edge of the colony to the edge of the clearing. => No ZOI means no antibiotic production => ZOI means antibiotic production **We didn't measure entire diameter of ZOI as we did in Kirby Bauer experiment because of the variability in the size and shape of the colonies.

Diazotrophs

Small amounts of nitrogen is fixed by purely physical processes like lightening and electricity. However, the bulk of nitrogen fixation is an enzyme driven process by diazotrophs (especially the symbiotic ones) ***Both free-living and symbiotic bacteria are capable of nitrogen fixation but the greatest contribution to the nitrogen cycle appears to be from the symbiotic species.

Antibiotic resistance

Some bacteria are naturally resistant to certain types of antibiotics. However, bacteria may also become resistant in two ways: 1) by acquiring resistance from another bacterium. or 2) by a genetic mutation **Different mutations yield different types of resistance: => some enable bacteria to produce enzymes that inactivate antibiotics => others eliminate the cell target of the antibiotic => others close up entry ports for antibiotic => others manufacturer pumping mechanisms to export antibiotic back outside so it doesn't reach its target

How is plasmolysis used in the food industry?

The dehydration of tissue (beef jerky) promotes the plasmolysis of any bacterial cells in the food and by preventing metabolic activity, controls the spoilage of the food. (many yeast and fungi are more tolerant of high osmotic pressure and may continue to grow on food treated in this manner)

Grid

The standard hemocytometer chamber contains one large square, which is comprised of 25 small squares. Each small square has 16 smallest squares. The volume of the one large square is 0.0001 ml Large square: => has 25 small squares Small square: => has 16 smallest squares **Using 40x objective lens under phase contrast setting, count the total number of organisms in 1 small square (total in 16 contiguous smallest squares) and multiply that by 25. Multiply by DF. And divide by the volume of 0.0001 ml. **The target number per smallest square is 5-15 cells so dilute if you need to. (# organisms in small square x 25 x DF)/0.0001 ml = #cells/ml

What are antibiotics?

They are SECONDARY METABOLITES - molecules produced by mircoorganisms that are not required directly for growth, but serve useful purposes for the microbes that make them, giving them an edge over those that do not. Examples of secondary metabolites: antibacterials, anticoccidials, antifungals antibiotics, etc. Definition of antibiotic (3): 1) produced by microorganisms 2) kills or prevents growth of other microorganisms 3) effective in small doses **antibiotics are not effective against the flu because flu is caused by a virus

Exponential/Log phase

This phase is called exponential phase because during each division, each cell divides into two and so the number of cells increases exponentially. The cells divide at a constant rate. The constant interval in which the cells divide is called the generation time. Generation/doubling time is calculated during exponential phase. (WHY? Because this is the most reproducible measure of the bacterial response to the environmental conditions.) **Because cell number (N) doubles with each division, the increase in cell number over time is exponential, not linear. Then how come in a bacterial growth curve we see the log phase being linear? Because we used a log scale for the y axis! **A linear increase on regular scaled y axis would only occur if the cell number rose by a FIXED amount after every generation (from 1 to 2, 2 to 3, 3 to 4, etc) But in exponential growth, the cell number doubles after every generation (from 1 to 2, 2 to 4, 4 to 8, etc)

Acidophiles

Thrives in acidic environments

Piezophiles

Thrives in high pressure like in hydrothermal vents in the deep sea.

Death phase

When number of cells decline exponentially due to lack of nutrients and accumulation of waste products. cell death > new cells **There are always a few unusually resistant cells that survive and can begin growth again when conditions are suitable.

Plasmolysis

[hypertonic solutions] When a cell is placed in a medium of HIGHER solute concentration, water out of the cell, resulting in shrinkage of cell and reduction in metabolic activity. If bacterial cell death occurs this condition is called bacteriostatic.

Plasmoptysis

[hypotonic solutions] When a cell is placed in a medium of LOWER solute concentration, water rushes into the cell and the cell ruptures. This is known as plasmoptysis. If bacterial cell death occurs this condition is called bacteriocidal.

Chemotherapeutic drugs

are compounds that are taken internally to ease the symptoms of a disease or to speed the patient's recovery. **Not all chemotherapeutic drugs are antibiotics. Chemotherapeutic drugs is called an antibiotic if it meets the 3 definitive critieria for antibiotics: 1) produced by microorganisms 2) kills or prevents growth of other microorganisms 3) effective in small doses

Streptococcus thermophilus

is a thermophil that can tolerate low pH and is one of two bacterial strains needed to make yogurt out of milk.

Stationary phase

Decline in nutrients and increase in waste products results in equilibrium between rate of cell deaths and cell division. (or could be because of a cessation of division altogether)

Which blank did we use? A) Water B) An empty tube C) Yeast cells D) Ethanol E) YPD broth

E) YPD broth **Always use the same medium that is used to grow the culture you are measuring because fluids will absorb light even in absence of cells. So having your zero baseline be the medium without the cells will give you an accurate indication of cell growth.

Microbes that produce antibiotics:

Fungi => genus Penicillin Bacteria => genus Bacillus Bacteria => genus Streptomyces **Streptomyces produces most medically important antibiotics **Streptomyces is part of group called Actinomycetes: => resemble fungi => gram + => Streptomyces produces Geosmin - earthly flavor and aroma, earthly taste of beets and scent of rain on dry eartch

Rhizobium and roots of legumes

***Nodulation only occurs if nitrate is limiting! Symbiosis (both live together), mutualistic relationship (both benefit): => Bacteria: Get protection and nutrients from the plant => Plant: Gets nitrogen from the bacteria in a form it can use

Legumes

***The PLANT secretes chemotactic flavenoids

Growth curve vs standard curve

**Both use spectrophotometer (The amount of light absorbed by a culture increases as the concentration of cells in the culture increases) **Both have OD values in Y axis. BUT growth curve uses a log scale for y axis. Standard curve doesn't. Growth curve has time in X axis where as standard curve has concentration in X axis. That is why their shapes are different. Growth curve has 4 phases, which gives the curves characteristic shape. Standard curve starts to level out flat at OD values above 0.8.

Nitrogenase is an oxygen sensitive enzyme

**Nitrogenase is an oxygen sensitive enzyme - requires anaerobic or extremely low oxygen conditions to be active ***Thus all nitrogen-fixing microbes develop strategies to keep oxygen levels low.

Rhizobium

**Rhizobia (NOT rhizobiUM) refers to bacteria that can form symbiotic relationships with legumes forming nodules **Many rhizobia is from the genus rhizobium. But not all species of rhizobium are called rhizobia (not all of them can form the characteristic symbiotic relationship that defines rhizobia) Rhizobium is: => gram negative rods => do not sporulate => motile => aerobic => a particular species of Rhizobium is only able to form nodules on certain legumes

Counting Viable Cells using Spread Plates

**Since each viable cell forms a colony, they are referred to as the colony forming units/ml (cfu/ml). In this experiment, we did a serial dilution of our culture and plated 100 ml of each dilution, then counted the cfu. Used that info to get cfu/ml of original culture. **A good countable plate has between 30-300 colonies. You plate different dilutions to hopefully end up with a plate that has colonies within that range. Calculating CFUs/ml of original culture => cfus/100 ml * CDF = cfu/ml of original culture **Benefits: -Only this method allows us to count viable cells. -It's able to measure very small number of cells per ml. **Disadvantages: -Takes longer because we have to wait at least 24 hours for colonies to form

You get on OD of 0.7 after dilution. Calculate the OD600 of the original culture.

**To go from OD of diluted to OD of undiluted (as in this example), you multiply OD of diluted by dilution factor. This example: OD of diluted = .7 Dilution factor = 3 OD of undiluted = OD of diluted*DF = .7 * 3 = 2.1 ****If you are ever asked for an OD of a different concentration, start with a concentration that will give you OD values between 0.1-0.8

You want to determine the concentration of yeast in a liquid culture. You measure the OD600, and find that it is 1.5, so you do a 1:3 dilution. What OD do you expect to read after the dilution? A) 1.5 B) 0.5 C) 0.75 D) 0.15 E) 0.375

**To go from OD of undiluted to OD of diluted (as in this example), you divide OD of undiluted by dilution factor. This example: OD of undiluted = 1.5 Dilution factor = 3 OD of diluted = OD of undiluted/DF = 1.5/3 = 0.5 ****The problem with this example is that the OD of undiluted is too high (above the accurate OD range) so it is inaccurate.

Temperature sensitivity

**Unlike mammals who are capable of homeostasis, the temperature of a microbial cell depends on the temperature of the environment Maximum growth temperature => highest temperature at which growth is possible Optimum growth temperature => the temperature at which growth is highest Minimum growth temperature => lowest temperature at which growth is possible

Heterocysts

**heterocysts have been modified in such a way as to render them semianaerobic **They are only produced when anabaena is growing in the absence of combined nitrogen

What can a spectrophotometer be used for?

1) Colorimetric assays to measure enzyme activity Note (from wikipedia): Colorimetric analysis is a method of determining the concentration of a chemical element or chemical compound in a solution with the aid of a color reagent 2) Determine concentration of DNA or RNA in solution 3) Measure the turbidity of a culture **Requires greater than 100,000 cells/ml **Benefits of using spectrophotometer is that it is quick and easy **Disadvantage is that it is not particularly precise and it doesn't differentiate between dead and alive cells

In a short sentence describe the 3 different methods of measuring microbial growth

1) Obtaining OD (optical density) as a measure of turbidity of a culture using a spectrophotometer 2) Counting the cells directly using a hemocytometer (the thick glass microscope slide) 3) Counting the number of viable cells by doing a colony count

The process of nodulation

1) Plant produces flavenoids that attract bacteria to specific sites on the root heairs. These flavenoids also activate transcription of bacterial Nod factor genes. => Rhizobia carry a Sym plasmid that encodes for: Nodulation genes Nitrogenase genes Host-specificity genes Auxin genes 2) Bacteria attach to recognition sites on root hair surface 3) Invagination of root hair membrane forms an infection thread. Bacteria are carried inside the infection thread. 4) Auxin genes are expressed (plant root growth regulating hormones). This stimulates root cells to divide, forming a nodule around the infection thread. 5) Bacteria continue to multiply 6) Within nodule, groups of 4-6 bacteriods are surrounded by plant membranes

Mesophiles

28C - 38C optimum growth temperature 20C-45C temp range

Which method will allow us to ONLY count live cells?

3) counting number of viable cells by doing a colony count **The others (spectrophotometer and hemocytometer) will not be able to distinguish between dead and alive bacteria.

Psychrohiles

< 16 C optimum growth temperature 4C-18C temp range

Bacterial growth curve

=> Growth is considered as increase of cell size and/or increase in cell numbers leading to an overall increase in biomass => We determine growth by measuring the number of cells at any given moment. This can be done using plate count technique or spectrophotometric method The difference? => plate count technique is able to measure a very small number of cells per ml => whereas the spectrophotometric method requires > 100,000 cells per ml but is easier to use

Differences between hemocytometer microscope slide and wet mount microscope slide

=> In the hemocytometer microscope slide, the volume of the grid is known and is exact. => The coverslip is ALWAYS placed on the grid before adding the culture.

How to kill mesophilic spoilage bacteria?

=> Many pathogenic mesophiles have an optimum temperature of 37C - normal human temp => Raising temp kill mesophilic spoilage bacteria **Preserving milk by converting it into yogurt requires the activity of high thermophilic bacterial strain Streptococcus thermophilus => Reducing temp in refrigerator prevents growth of spoilage bacteria **However phsychrophilic spoilage bacteria do exist. This accounts for the fact that food can still spoil in the refrigerator!

Rhizobia carry a Sym plasmid that encodes for:

=> Nodulation genes => Nitrogenase genes => Host-specificity genes => Auxin genes

Summary of different ways food can be protected from spoilage bacteria

=> Temperature to kill mesophilic spoilage bacteria => Drying food to promote plasmolysis to prevent microbe metabolic activity => Fermenting lactose in milk to decrease pH and kill spoilage bacteria Ex: Streptococcus thermophilus Ex: Lactobacillus bulgaricus **The two strains required to make yogurt from milk. Both are resistant to low pH.

How are heterocysts different from vegetative anabaena cells?

=> Three additional cell walls including one of glycolipid that forms barrier to oxygen => Expresses nitrogenase and other proteins involved in nitrogen fixation => Degrades photosystem II, which produces oxygen => less photosynthesis => Maintains photosystem I, which allows them to generate ATP by cyclic photophosphorylation **Where do heterocysts get carbohydrates from? => From the vegetative cells around them => Fixed carbon and nitrogen sources are exchanged through channels between the cells

Anabaena protects the function of nitrogenase by:

=> Using anaerobic respiration for energy-production (it degrades photosystem II but maintains photosystem I) (see previous flashcard) => Produces specialized, semi-anaerobic cells for nitrogen fixation (Heterocysts)

- Explain a zone of inhibition. What determines its size?

=> after isolating the pathogen, it is heavily streaked on Mueller-Hinton agar plate => antibiotic discs are placed. and plate is incubated. **During incubation, the antibiotic diffuses from the disk into the agar, creating a concentration gradient such that the greatest concentration is at the edge of the disc

What does the generation time/doubling time depend on?

=> bacterial species => type of medium => temperature => pH **Exponential growth occurs only for a short period when all nutrients are in full supply and the concentration of waste products has not become a limiting factor.

What would you target to kill a bacterial pathogen?

=> cell membrane (Colistin, Polymyxin) => cell wall (penicillin, all the "-cillins") => metabolic activity (Sulfonamides) => DNA replication (Ciprofloxacin, Rifampicin) => nutrient uptake => protein synthesis (kanamycin)

- Vibrio natriegens growth curve

=> has the fastest known doubling times measured under ideal conditions => a microbe found in salt marshes **In this experiment, we made four tubes (2 controls and 2 tests). The test tubes had added an antibiotic when the OD600 reached ~0.4. The OD values were measured every 10 min. The broth used was BHI (Brain Heart Infusion) - 2% NaCl

What is the size of ZOI affected by?

=> how easily the antibiotics diffuses in agar => type of medium used => inoculum size => incubation conditions (time, temp, etc) **it is important to keep as many of those factors constant so that we can relate the inhibition-zone size to whether the isolated microbe is R, S, or I in it's response to a particular antibiotic ^^^This is the type of wording you need on the exam **the Kirby-Bauer method is highly standardized to avoid varying results due to alterations in procedure ****IF THE KIRBY BAUER METHOD IS USED to standardize the conditions, then the size of ZOI is related to two things: 1) MIC - (minimum inhibitory concentration) minimum concentration that kills or inhibits that microbe 2) ability of antibiotic to diffuse through the agar The size of the zone that indicates sensitivity to an antibiotic varies greatly per antibiotic and per isolate. These differences reflect the different concentration gradients formed by the different antibiotics during diffusion through the agar and under the standard conditions of Kirby-Bauer method, the different positions of the MICs on the gradients.

Strategies to keep oxygen levels low in presence of nitrogenase (3)

=> rapid respiration of available oxygen before nitrogenase starts functioning => sequestering of the microbes (in symbiotic bacteria) => use of protective compounds to reduce the amount of available oxygen

Antibiotics vs antibacterials

Antibiotics are more board and can act on fungi, bacteria, and other compounds. Antibacterials are a specific type of antibiotics that act only on bacteria.

Temperature preferences of different domains

Archaea has the greatest degree of tolerance to heat (lower degree of temp sensitivity) Bacteria has inbetween degree of tolerance to heat (inbetween degree of temp sensitivity) Eukarya has the lowest degree of tolerance to heat (higher degree of temp sensitivity)

- Under what circumstances would a clinical lab investigate a pathogen further to see which antibiotics have an effect on it?

A patient has an infection and goes to the doctor. The doctor can make a good guess to what causes the infection. If antibiotics are needed, they will traditionally prescribe broad-spectrum antibiotics. If the patient starts recovering within 24-48 hours, no further investigation is needed. If the infection appears chronic (lasts a long time or occurs frequently) then the pathogen is often isolated, identified and tested for its susceptibility to various antibiotics, in order to decide which chemotherapeutic drug to prescribe next. The microbiologist who will do this job, will probably use the Kirby-Bauer method. **if infection appears chronic even after being prescribed a broad-spectrum antibiotic, the pathogen will be further investigated to see which various antibiotics is the pathogen susceptible to in order to decide the best chemotherapeutic drug to prescribe next. They will do this using the KIRBY-BAUER METHOD!

The spectrophotometer can only accurately measure OD values between? A) 0.1-0.8 B) 0.1-0.9 C) 0.2-1.0 D) 0.0-1.0 E) 0.5-0.8

A) OD values between 0.1 and 0.8 This is because OD < 0.1 may not remain steady and OD > 0.8 are not linear

Anabaena is a: A) photoautotroph B) photoheterotroph C) chemoautotroph D) chemoheterotroph E) space alien

A) photoautotroph ***photo/chemo refers to energy source ***auto/hetero refers to organic molecule source Autotroph - produces complex organic molecules by themselves => photoautotroph - can fix CO2 using light as their energy source (photosynthesis) => chemoautotroph - can fix CO2 using INORGANIC energy sources Heterotroph - cannot produce complex organic molecules by themselves and have to consume it from the environment => photoheterotroph - uses light as energy source => chemotheterotroph - uses INORGANIC energy sources

What is the correct order of a bacterial growth curve? A) Log phase, lag phase, stationary phase, death phase B) Stationary phase, log phase, death phase, lag phase C) Lag phase, log phase, stationary phase, death phase D) Death phase, lag phase, log phase, stationary phase E) Lag phase, stationary phase, log phase, death phase

C) Lag phase, log phase, stationary phase, death phase

What extremes are we testing in our experiment? A) Temperature, oxygen and salt concentration B) pH, pressure and temperature C) pH, salt concentration and temperature D) Salt concentration, sugar concentration and pH E) Salt concentration, pH and levels of nutrient

C) pH, salt concentration and temperature

Nonhalophile

Cannot grow under saline conditions