Chapter 6- Microbial Growth Flashcards
purple/green sulfur bacteria
photoautotrophs that are anoxygenic that use light as a primary source of energy, but then use sulfur to convert CO2 to sugar
purple/green nonsulfur bacteria
Use light as primary source of energy, but use organic compounds to reduce CO2.
What distinguishes purple/green sulfur bacteria from each other?
green- chlorophyll located in vesicles (chlorosomes) underlying and attached to plasma membrane
purple- located in invaginations (chromatophores).
location of stored sulfur
16S rRNA sequence
Harmful Oxygen species
Singlet- O2- excited form of O2 that has an e- in a higher orbital due to energy absorption
superoxide free radicals- O2^1- : formed from respiration; can be neutralized to oxygen and hydrogen peroxide by superoxide dismutase in organisms that can at least tolerate oxygenated conditions
peroxide anion- O2^2-: comes from hydrogen peroxide. catalase can convert hydrogen peroxide into water and oxygen, while peroxidase can convert it to water. produced from respiration
hydroxyl radical- OH*: formed by ionizing radiation and respiration in traces; THE MOST REACTIVE
Which organisms have catalase/ superoxide dismutase
Obligate aerobe - both Facultative anaerobe - both Obligate anaerobe - neither Aerotolerant anaerobe - only SOD Microaerophiles - SOD+ catalase +/- (small amounts
Biofilm characteristics
3-D structure not well appreciated until the development of confocal microscopy
usually attached to a surface, may facilitate conjugation, works cooperatively, sheltered from desiccation, antibiotics and the immune system, 10-1000x’s resistant to antimicrobials.
Quorum sensing
the ability of bacteria to coordinate gene expression with other bacteria using signaling molecules; usually respond to local population density. coordinates genes for surfactants, individual/group growth, and biofilm dispersion.
How can you prevent biofilm formation?
Block chemical signals, incoporate antimicrobials into surfaces, use lactoferrin to stimulate surface motility and prevent formation.
Soft agar and solid agar
Soft agar: less than a 1% agar solution, used to test for motility
solid agar: 1.5-1.8%, regular culture
Ways to remove O2 from media prior to inoculation?
heating- liquid media can be heated to drive-off dissolved O2 then you can seal it.
reducing media- using a chemical like thioglycolate reduces O2 to water.
incubating a plate in an oxygen free environment (anaerobic jar with bicarbonate, borohydride and Pd catalyst pellets) *this is useful if brief exposure to oxygen isn’t fatal.
incorporating oxyrase into media (useful if brief O2 exposure isn’t lethal).
Capnophiles
Grow better at high CO2 concentrations (because of the similarity to the intestines, respiratory tract, and other tissues)
Dextrose agar
selective media that encourages growth of fungi by acidic pH of 5.6 and the presence of chloramphenicol, which is an antibiotic that targets prokaryotes.
Bismuth agar
selective/differential medium that differentiates Salmonella species, usually S. typhi from feces. Bismuth inhibits gram + bacteria and most gram - bacteria except Salmonella.
*broth also contains ferrous sulfate because S.typhi is the only Salmonella species that reduces it to hydrogen sulfide. *
Blood agar
distinguishes bacteria that lyse RBCs (ex. S. pyogenes)
mannitol salt agar
used to select for/distinguish S.aureus
has high salt 7.5% NaCl to discourage growth of other organisms, and has a pH indicator that changes color when mannitol is fermented.
MacConkey Agar
distinguishes and selects for Gram - cells that ferment lactose.
Bile salts and CV discourage growth of Gram +, lactose + pH indicator are for lactose fementers that will produce pink or red colonies.
Enrichment culture
Used to favor the growth of an organism, designed to increase very small levels to detectable levels (usually done in liquid and for fecal/soil samples). Example: boiling a culture to get Clostridium endospores, or isolating for a bacterium that can metabolize phenol.
preserving bacterial cultures
days-weeks: refrigeration
years:
- deep freezing -80 C or - lyophilization (-54 to -72 C ) then sublimated in a vacuum, when lyophilization is complete, ampule neck sealed under vacuum.
Methods of bacterial division
- binary fission (mainly)
- budding (plantomycetes, cyanobacteria, firmicutes)
- fragmentation (few filamentous bacteria)
Lag Phase
Period of adjustment to new conditions where little to no division occurs (pop doesn’t increase); metabolic activity is high and this process may last 1hr-few days
Log phase
generation time reaches a constant minimum, the number of cells produced greatly exceeds the number of cells dying, and the metabolic activity is at its highest; cells are most susceptible to adverse factors such as radiation or antibiotics.
stationary phase
pop begins to stay static (cells producing = cells dying), cell number doesn’t increase, cell division slows down. Factors that can slow down microbial growth is the accumulation of toxins, acidic pH, limited nutrients or insufficient O2
Death phase
population size decreases (cells dying > cells producing), cell number decreases logarithmically, cells lose ability to divide, and a few cells may remain alive for a long period of time.
plate count
(DIRECT) spread plate with a sample and count number of colonies (assumptions are that each colony originates from a single cell and each inoculum is homogenous; the former isn’t always true). it measures only viable cells, but it takes 24hrs+, and only counts between 25-250 colonies are accurate. There are also plate counts, where you can add more culture or determine aerotolerance, but heat sensitivity could be a problem, embedded colonies may not be transferable, some differential plates require surface colonies for morphology diagnoses.
Filtration
(DIRECT) used to measure dilute bacterial samples. first a sample is filtered and then the filter is transferred onto a petri dish to then incubate and then count the colonies.
Most probable number
(DIRECT) used to measure bacteria that will not grow on solid medium. Make several dilutions of a culture and inoculate a broth tube for each one, then count the number of positive tubes in each set.
Direct Microscopic count
bacteria/mL = # cells counted/ volume of area counted
(DIRECT) a bacterial susmension (0.01mL) is placed on a slide with a grid and the cells are counted and multiplied by a volume factor to obtain concentration. there is no incubation time required, but you cannot always distinguish between live and dead cells, motile bacteria are difficult to count, and it requires a high concentration of bacteria (10mil/mL).
Turbidity
(INDIRECT) as bacteria multiply in a broth tube, the media becomes more turbid. there is no incubation time required, but you cannot distinguish between live and dead cells and it requires a high concentration of cells (10-100mil cells/mL)
%transmittance correlates to cells/mL; you plot the OD units on a graph against viable counts from plate counts (standard curve).
Absorbance/optical density = 2 - log % Transmittance
Metabolic activity
(INDIRECT) assumes the amount of metabolic product is in direct proportion to cell number.
Dry weight
(INDIRECT) best for measuring filamentous bacteria or fungi. cells are centrifuged to then be desiccated and weighed. Wet weight and volume of cells after centrifugation measurements can be made. However, it does’t distinguish live and dead cells.
