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Microbiology 2 2


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minimal inhibitory concentration
lowest dilution with no turbidity.
Minimal bactericidal concentration (MBC)
lowest dilution With no growth when subbed.
Factors that influence disk diffusion susceptibility in bauer kirby?
medium, agar depth, disk potency, inoculum concentration, pH, and β-lactamase production by the test organisms.
Inhibition of cell wall synthesis
penicillin, vancomycin, bacitracin, penicillins, cephalosporins, monobactams, carbapenems
Combination with cell membranes
polymixin causes holes in cytoplasm
Action on eukaryotic membranes
the polyenes: sterols present in the cell wall of eukaryotes. Fungi are inhibited, but dangerous for human use, as it is not as selectively toxic.
Inhibition of protein synthesis
chloramphenicol, streptomycin, neomycin, kanamycin, tetracyclines, erythromycin
50S ribosome subunits
erythromycin (macrolides), chloramphenicol, clindamycin, lincomycin
30S ribosome subunits
30S ribosome subunits
streptomycin, gentamicin, tobramycin, kanamicin, amikacin.
(urinary tract only, not antibiotic)
DNA gyrase
nalidixic acid, norfloxacin, ciprofloxacin, ofloxacin (quinolones) Novobiocin
DNA directed RNA polymerase
compound produced by bacteria active against closely related bacteria.
Ciprofloxacin is used to treat urinary tract infections and anthrax caused by penicillin-resistant bacillus anthracis.
Fluorinated derivatives of nalidixic acid are more soluble than nalidixic acid and reach clinically therapeutic levels in blood and tissues.
It is broken down in the body to sulfanilamide, the active agent
Sulfanilamide competes with _________ which is a precursor of __________
para aminobenzoic acid
folic acid
Each year more than _____ metric tons of chemotherapeutic agents are manufactured.
Mechanisms of antibiotic Resistance?
1. Resistance may be inherent or can be acquired.
2. Organism may not have the structure affected. Ex. Mycoplasma have no cell wall - are resistant to penicillin.
3. Organism may be impermeable to antibiotic.
4. Organism alters the antibiotic to an inactive form.
5. Organism may modify the target of the antibiotic.
6. By genetic change, the organism may alter the metabolic pathway that the antibiotic blocks.
7. The organism may be able to pump out any antibiotic entering the cell (efflux).
8. Genetic change: due to plasmid transfer or at the chromosomal level.
Type of resistance:
inactivation of antibiotic, e.g. penicilinase; methylases, acetylases phosphoryl-ases
penicillins, chlorampheni-col
plasmid and chromosomal
Type of resistance:
reduced permeability
Type of resistance:
tetracyclines, chloramph.
plasmid chromosomal
Type of resistance:
alteration of target; ribos.
Type of resistance:
alteration of target; DNA gyrase
Type of resistance:
development of resistant pathway
Polyenes - disrupt membrane integrity
polyoxins - inhbit chitin synthesis
azoles and allyamines - inhibit ergosterol synthesis
5-fluorcytosine - nucleotide analog that inhibits na synthesis
griseofulvin - inhibits microtubule formation
complete virus particle

complete pacakge is called?
protein coat surrounding nucleic acid center.

protein subunit making up capsid
Viral structure
technically geometrical structures called icosahedrons with 20 triangular faces and 12 corners or rod like. Some are enveloped. Roughly 30-300 nm in size.
Viral quantification:
Phage overlay
plaques due to lysis, pfu.
Viral quantification:
Animal viruses
tissue culture, cpe, count analogous to plaques.
Viral Replication
(adsorption) of virion to susceptible host. High specificity for host.
Viral Replication
Penetration (injection
infectivity of the virus particles disappears, called eclipse, due to the uncoating of the virus particles
Viral Replication
Early steps in replication
host cell biosynthetic machinery is altered as prelude to virus nucleic acid synthesis, virus specific enzymes are typically made.
Viral Replication
Latent Period
Replication of virus nucleic acid and protein. No infectious virions are present extracellularly.
Viral Replication
Assembly of structural subunits
packaging of nucleic acid into new virus particles. Titer of active virions inside cell rises dramatically. At this time, if the cells are broken open, active virus can be detected. The protein coat forms spontaneously from protein capsomeres by self assembly.
Viral Replication
Release of mature virions
due to cell lysis, budding or excretion process. Release of virions occurs after breakdown of cw by lysozyme like protein.
Burst size
the no. Of virions released, varies with virus and host cell.
infectivity of the virus particles disappears
Viral Replication
varies from 20-60 min in bacteria. Viruses to 8-40hr in most animal viruses.
Early proteins
synthesized soon after infection, necessary for replication of viral nucleic acid
Late proteins
synthesized later, protein coat.
Temperate bacteriophages
Direct to lysis, or integration of the virus DNA into the host DNA – lysogenization. The lysogenic cell can also be induced to produce mature virus and lyse.
Main Types of bacterial viruses
RNA ss
MS2 (small globular)
Main Types of bacterial viruses
RNA ds
q6 (Small globular,membranous)
Main Types of bacterial viruses
DNA ss
T3,T7 (Small lander, no neck)
Main Types of bacterial viruses
DNA ds
T2, 74 (Large lander, long, coiled neck)
Class I
ds DNA +/-
Class II
ss DNA + or -
dsDNA +/-
Class III
ds RNA
Class IV
ss RNA +
Virus genome serves directly as mRNA.
Class V
ss RNA –
RNA viruses, whether +, - or ds require a _________.
specific RNA -dependent RNA polymerase.
RNA Replicase.
virus specific and RNA-dependent RNA polymerase
Class VI
ss RNA +
Process of copying information from RNA into DNA is called ____, requiring an enzyme called _____.
reverse transcription
reverse transcriptase
- triplet of bases in mRNA that code for an amino acid.
triplet of bases in tRNA that code for an amino acid.
A single amino acid is frequently coded for by several different but related triplets of bases.
Nonsense codons
Those that don’t lead to amino acid production. May function as punctuation - ie termination of the growing polypeptide chain.
lateral gene flow
genes are transferred from donor to recipient, rather than mother to daughter.
the genotype of an organism is designated by
3 lowercase letters followed by a capital letter all in italics indicating the gene involved, e.g. hisC
The phenotype of an organism is designated by
a capital followed by 2 lowercase and a + or – to indicate presence or absence of property.
E.g. His+ strain is capable of making histidine, whereas his- is incapable.
selectable mutation
gives the mutant a clear advantage under certain environmental conditions
Lederberg replica plating technique
used for screening nutritional mutants.
a mutant that has a nutritional requirement for growth.
parent from which auxotroph is derived.
Bulk of spontaneous mutations occur during DNA replication as result of
errors in pairing bases.
Point mutations
Mutations involving a change in one base pair
silent mutations
Because of degeneracy, not all mutations in polypeptide encoding genes result in changes in polypeptide
missense mutation
Changes in 1st or 2nd base more often lead to significant changes in peptide leading to an inactivity or reduced activity.
nonsense mutation
point mutation resulting in termination of translation causing incomplete polypeptide due to formation of stop codon.
substitution of one purine base (a or g) for another purine , or one pyrimidine (c or t) is substituted for another pyrimidine.
point mutation where a purine base is substituted for a pyrimidine or vice versa.
Frameshift mutation
deletion or insertion of a base pair causes this with serious results. This mutation is a result of replication error. Shifts in the reading frame of messenger RNA are caused by insertion or deletion mutations in DNA.
Transposon mutagenesis
insertion of a transposable element occurs within a gene, loss of gene function results. Transposable elements can enter chromosome at various locations and are used by microbial geneticists as mutagenic agents.
Back mutations or reversions
point mutations are reversible by process of reversion, resulting in a revertant, a strain in which the wild type phenotype that was lost in the mutant is restored. Can be due to restoration at the same site or occur at a different site (called suppressor mutation).
chemical, physical, or biological agents to increase the mutation rate
Nucleotide base analogs
molecules resembling DNA purine and pyrimidine bases in structure but with faulty pairing properties.
can base pair with guanine causing at to gc substitutions.
can base pair with cytosine, causing at to gc substitutions.
Alkylating agents
powerful mutagens that induce at higher frequency than analogs.
Nitrous acid
Deaminates a & c
Reacts with c
Put methyl on g
Cross link DNA strands
Both point mutation and deletions
Intercalative dyes
Acridines, ethidium bromide
Inserts between 2 base pairs
Microinsertions and microdeletions
Pyrimidine dimer
Repair may lead to error or deletion
Free radical, break chain
Repair may lead to error or deletion
1. Transformation
free DNA inserted directly into competent recipient cell.
2. Transduction
transfer of bacterial DNA via temperate or defective phage.
3. Conjugation
DNA transfer via actual cell to cell contact between donor and recipient.
the ability of the recipient to take up free DNA. Only certain strains are competent and this seems to be genetically determined.
In bacillus only about ____ of the cells become transformable while in streptococcus, ____ but for only a few minutes.
In ___ only about 20% of the cells become transformable while in ____, 100% but for only a few minutes.
DNA is bound at cell surface by a DNA binding protein, after which the entire double stranded fragment is taken up, or a nuclease degrades one and the other is taken up.
the bacteria transformed with DNA extracted from a bacterial virus. It has become a standard mechanism for studying transformation.
Competence can be induced in non-transforming bacteria by ____
treatment with calcium in the cold.
Generalized transduction
host DNA from any portion of host genome becomes part of mature virus in place of virus genome. If the donor genes don’t undergo homologous recombination with host genome, they will be lost, as they can’t replicate independently and are not part of virus genome.
Specialized transduction
only in some temperate viruses; DNA from a specific region of the host genome is integrated directly in to virus genome, usually replacing some virus genes. The virus is in both cases is usually non-infective (therefore called defective) as a virus because bacterial genes have replaced some necessary viral genes.
transducing particle
When infected with phage, the lytic cycle may be initiated. The enzymes responsible for packaging viral DNA, sometimes package host DNA by accident.
The resulting virion is called a transducing particle. When released, the lysate contains mixture of normal and transducing particles.
conjugative plasmids
A process by which plasmids are transferred from donor to recipient [cell to cell transfer]. This is encoded by some plasmids themselves
Some plasmids called episomes can integrate into the chromosome, and under these conditions their replication comes under control of the chromosome. This is similar to that of several viruses whose genome can become incorporated prophages.
Plasmids can be eliminated from cells (curing) by various treatments. This causes inhibition of plasmid replication without inhibition of chromosome replication.
Transmissibility by conjugation is controlled by set of genes within plasmid called the ____ region
rolling circle replication
Process called rolling circle replication best explains DNA transfer. 1 strand of plasmid DNA circle is nicked and is transferred to recipient. The enzyme also has helicase activity and is involved in unwinding the strand to be transferred.
As this transfer occurs, DNA synthesis by rolling circle mechanism replaces the transferred strand in donor with complementary strand which is also then made in recipient.
– if plasmid acts like an episome and integrates with host chromosome, then chromosomal genes are mobilized and transfer of host genes takes place.
Those that have a chromosome integrated f plasmid are called hfr and are unable to take up a 2nd f plasmid.
Metabolically related populations of microorganisms
Energy enters ecosystems in form of
sunlight, organic carbon, and reduced inorganic substances.
allochthonous organic matter
organic matter that enters from the outside
Biogeochemical cycle
microorganisms play a role in the recycling of several elements , the study of which is called biogeochemistry.
prime niche
Ecological theory states that for each organism there is a prime niche.
Small organisms live in small environments so they are called microenvironments. E.g. A 3 mm soil particle can contain several different microenvironments, different chemically and physically.
Experiments demonstrate their oxygen content can vary, even throughout such a small particle with the outer zones being fully ____, and the inner zones completely _____ with intermediate or microaerophilic zones between.
Organisms sticking to surface, encased in slime, usually polysaccharide. Biofilms trap nutrients essential for growth. Cell – cell communication is critical in biofilm development and maintenance. Attachment is a signal for the expression of biofilm specific genes, which encode proteins that synthesize cell to cell signaling molecules that begin polysaccharide formation.
homoserine lactones
The major signaling molecules in pseudomonas biofilm formation
quorum sensing
chemotactic agents to recruit nearby organisms,
Reasons for biofilm formation
1- Allows cells to remain in favorable niche.
2- Allows cells to live in close association fostering genetic exchange and quorum sensing.
3- Type of defense – resist physical force of current or phagocytosis.
4- Biofilms may be the typical way bacteria grow in nature.
mineral soils
weathering of rock
organic soils
sedimentation from bogs or marshes
soil around roots
Predominant microorganisms in oxic areas
cyanobacteria and algae
Predominant microorganisms in anoxic areas
anoxygenic phototrophs
algae floating or suspended in water
Benthic algae
those attached to bottom or sides.
Adapted to grow under dilute conditions.
warmer and less dense surface layer in lakes
cold dense bottom layers in lakes
zone of transition between these two layers in lakes
Biochemical oxygen demand (bod)-
Oxygen consuming property of a body of water – directly related to amount of suspended organic matter.
Objective in water treatment
reduce bod.
often develop in response to pollution by inorganic nutrients
Algae, cyanobacteria, and macrophytes (aquatic weeds)
The largest carbon reservoir is
earth’s crust
global turnover time of carbon
about 40 years
complex mixture from protoplasmic constituents of soil microorganisms.
most rapid turnover of carbon
Co2 of atmosphere
The single most important contribution of co2 to the atmosphere
microbial decomposition of dead organic material.
The only way that new organic c is synthesized
photosynthesis (more important), chemosynthesis (less important).
make methane
consume methane
Methanogenesis is carried out by a group of
methanogens, strict anaerobes
Most methanogens use ___ as a terminal electron acceptor in ____ respiration, reducing it to ____ with h2.
c02, anaerobic, methane
primary fermenters.
decompose complex molecules
a species that lives off another species
secondary fermenters
species that ferment the products of primary fermentors
Acetate and h2 + co2 from primary fermentations can be directly converted to methane, although h2 + co2 can also be consumed by them.
Primary treatment of wastewater
Wastewater is pumped into the reservoir (left) where settling of solids occurs. As the water level rises, the water spills through the grates to successively lower levels. Water at the lowest level, now free of solids, enters the spillway (arrow) and is pumped to a secondary treatment facility.
Anoxic secondary wastewater treatment processes
Anoxic sludge digester
Trickling filter
Aeration tank
Activated sludge installation
fungal body
unit of structure of a mold, a tube shaped structure exhibiting cytoplasmic streaming toward tip as it grows
– hyphae forming an interwoven mat of growth consisting of vegetative hyphae extending into medium and aerial hyphae extending into the air above medium.
root like structure
description of hypha, a tube like structure with many nuclei.
asexual spore borne at the end of a conidophore.
sack like structure containing sporangiospores (asexual).
specialized sex hypha that grow toward each other.
sexual spores borne in a sack like structure the ascus.
spores borne on a structure called a basidium.
the fructification holding many basidia.
types of fungi
1- Ascomycetes
2- Zygomycetes
3- Basidiomycetes
4- Deuteromycetes
Aerial hyphae
extend above surface
Vegetative hyphae
below surface
Aseptate hyphae
no cross walls
Septate hyphae
cross walls
Budding yeast
no hyphae
Asexual Fungi
fungi imperfecti
Sexual fungi
Traditional biotechnology
large scale manufacture by microorganisms of products that they normally make.
Engineered organisms or recombinant DNA
use of organisms with modified genome specifically selected and placed into organism.
refers to an organism that ferments
to a vessel that is used for an industrial microbial process; it may be either aerobic or anaerobic
Metabolic specialists
microbes that have been altered by mutation to produce large amounts of metabolites.
Strain improvement
eg penicillium chrysogenum, 1µl/ml vs 85000 µl/ml –achieved via mutation and selection.
Ideal industrial microorganisms:
a. Grow rapidly – doesn’t tie up equipment, contamination less likely, easier to control environment
b. Are non-pathogenic
c. Are free of toxins
d. Can be removed from culture easily, e.g. Yeasts, filamentous fungi, filamentous bacteria
cells are used to chemically convert a specific substance from one form to another.
Microbial products
a. Cells themselves, e.g. Yeasts, lactobacillus, mushrooms.
b. Enzymes – amylases, proteases, lipases, rennin, isomerase (fructose from glucose), penicillin acylase (used in production of semisynthetic penicillins).
c. Commodity metabolites – ethanol, acetic acid, lactic acid, citric acid, methanol.
d. Growth factors – vitamins, amino acids, antibiotics, steroids, alkaloids
use of microbial products:
antibiotics, steroids, steroid conversion, hormones.
use of microbial products:
Genetically engineered
insulin, human growth hormone, antiviral, anti-tumor, e.g. Interferons, lymphokines, blood clotting, vaccines (hbv), monoclonal antibodies.
use of microbial products:
Agricultural uses
insulin, human growth hormone, antiviral, anti-tumor, e.g. Interferons, lymphokines, blood clotting, vaccines (hbv), monoclonal antibodies.
use of microbial products:
Specialty chemicals and food additives
amino acids, glutamic acid, lysine, tryptophan, aspartic acid, aspartame, phenylalanine, vitamins.
use of microbial products:
Commodity chemicals
energy production.
Secondary Metabolites
growth phase.
Secondary Metabolites
metabolic production phase.
fermentor probe
computer converts these data into microbiological or biochemical terms, and graphs the data. Processes data to measure growth, decides when and how much nutrient to add, optimal conditions maintained.
β-lactam antibiotics
Penicillins and cephalosporins, medically useful, contain the β-lactam ring, a complex 2 membered heterocyclic ring. They act by inhibiting peptioglycan synthesis.
Major ingredient of penicillin production is
corn steep liquor
Cyanocobalamin (B12)
synthesized in nature exclusively by microorganisms. B12 has cobalt as a central part of its structure; yields are increased by adding cobalt to the medium.

Propionibacterium yield from 19 – 23 mg/l, pseudomonas denitrificans produceds 60 mg/l in a 2 stage process using molasses as the c source.
Riboflavin (B2)
synthesized by many organisms including bacteria, yeasts and fungi. Ashbya gossypii produces up to 7g/l
Glutamic acid
used as a flavor enhancer. (MSG)
Aspartic acid and Phenylalanine
the ingredients of Aspartame.
an essential amino acid and is produced by brevibacterium flavum for use as a food additive.
steroid hormones created in microbial bioconversion
Induced enzymes
inducer is present, therfore added during stationary phase.
bacterial proteases
used in laundry detergents that also include amylases, lipases, reductases.
Laundry enzymes
formed by alkaliphilic bacteria, mainly bacillus licheniformis.
Amylases and glucoamylases
are important in the production of glucose from starch that can then be isomerized to fructose used in the food industry as high fructose syrup, made from corn, wheat, potatoes.
enzymes made by hyperthermophilic microorganisms, function at high temperature. Useful because many industrial processes work best at high temperature.

Ex- Taq and pfu DNA polymerases used in pcr (polymerase chain reaction).
Thermostable ___, ____ and ____ isolated and characterized.

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