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GVSU / Biology / BIOL 212 / How does the tryptophan operon work?

How does the tryptophan operon work?

How does the tryptophan operon work?

Description

School: Grand Valley State University
Department: Biology
Course: Introduction to Microbiology
Term: Summer 2015
Tags: antibiotics, Microbial, and control
Cost: 50
Name: Microbiology Unit 3 Study Guide
Description: These notes cover the notes I didn't post from Unit 2 and the notes needed for our third exam (Unit 3). They are a combination of notes from class and the reading. Things that are highlighted in a light gray are things I was unsure about (there are only two little things I think).
Uploaded: 03/20/2017
36 Pages 35 Views 7 Unlocks
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Unit 3 2/19/17 7:00 PM


How does the tryptophan operon work?



Repressible Operon: Operon that is always active unless repressed • Tryptophan Operon

o No Tryptophan = inactive repressor and transcription of  

operon continues

o Tryptophan present = tryptophan binds to the repressor and  activates it, causing it to bind to the operon and block  

transcription

o Inside the RNA transcript there is a leader region with two  

Tryptophan codons that will need to be translated before  

structural genes are translated, keeping this in mind…

▪ If Tryptophan is LOW there isn’t many tRNA charged  

with Trp so as transcription moves translation/the  

ribosome is close behind but when it gets to the leader  


What is an example of a spontaneous mutation?



If you want to learn more check out What event led to the making of the code of ethics?

region with 2 Trp codons it stalls because there is not a  

lot of charged tRNA. The RNA polymerase/transcription  

gets ahead and before the ribosome can catch up to  

translate it the mRNA #2 segment and the #3 segment  

bind together. This loop is far enough away from where  

transcription is happening that it doesn’t cause tension  

and transcription can continue.  

???? Trp Low = 2-3 bind = Transcription continue

▪ If Tryptophan is HIGH transcription is proceeding and  

translation/ribosome is close behind, there is enough  

tryptophan charged tRNA that the ribosome gets  Don't forget about the age old question of What is an example of inoculation?

through the Trp part of the leader region fine and  


What are the three major mechanisms of dna repair?



instead of 2-3 binding segment #3 and #4 bind  

together. This creates a termination loop close to where  

transcription is occurring that causes tension on the  

transcript and halts transcription.

???? Trp High = 3-4 bind = Transcription stops  

Mutations: Change in phenotype due to change in genotype (nucleotide  base sequence)  

• Wild-type: natural, non-mutated characteristic

• Mutant: organism with a mutation, its morphology, nutritional  characteristics, genetic control mechanisms, resistance to chemicals  and more vary from the norm.  

• Causes of Mutations

o Spontaneous Mutations: Random change in the DNA  sequence that happens without a known cause  

o Induced Mutations: change in the DNA sequence that is  caused by exposure to known mutagens, either physical or  chemical agents (Chemicals, nitrogen base analogs, radiation)  • Categories of Mutations  Don't forget about the age old question of What are examples of cognitive appraisal?

o Substitution Mutations

▪ Missense Mutation: change in a base pair that results in  a codon that codes for a different amino acid in  

translation  

▪ Nonsense Mutation: change in a base pair that results  in a stop codon and early termination of the polypeptide  during translation We also discuss several other topics like Why is self-defense justified?

▪ Silent Mutation: change in a base pair that changes the  codon but still codes for the same amino acid due to the  redundancy of the genetic code  

o Back Mutation: Reversion, when a mutated gene reverses  back to its original base composition

o Frameshift Mutation: insertion or deletion of a single base  that causes the nucleotides after the mutation to shift  (reading frame of mRNA shifts) resulting in change in codons  and different polypeptides  

▪ It is a given that stop codons will be produced  

downstream in this form of mutation, this happens  

whether the bases are added or deleted

• Mutation Repair Mechanisms

o Light Repair: When a thymine dimer (thymines that are right  next to each other bind together) is created it causes a  mutation in the strand when the strands have to replicate, in  light repair the enzyme photolyase gets energy from the sun  and breaks the bonds of the dimers to reverse the strand to  its normal DNA sequence.Don't forget about the age old question of Who won the sino japanese war?

o Base-excision Repair: enzymes (Glycosylases) remove a  section of the DNA that has an error in it and DNA polymerase  I and ligase fills in the gap  

o Nucleotide Excision Repair/Dark Repair: enzymes cut out a  short section of the DNA strand (about 12 nucleotides) that  contains a dimer and DNA polymerase I and DNA ligase repair  the gap with help of the undamaged complementary strand  

o Mismatch Repair: enzymes scan newly synthesized DNA  strands (they can tell which ones are new and which are old  because the new are non-methylated for a short period of  time) for mismatched bases and remove and replace them

o SOS Response: Prokaryotic cells with a lot of DNA damage  use a variety of processes to induce DNA polymerase to  

replicate the damaged strand, replication of this strand may  introduce new and possibly fatal mutations but it does allow a  few offspring to survive.  

*THIS IS THE BEGINNING OF MATERIAL FOR EXAM 3*

• Effects of Mutations

o Cause nonfunctional proteins = could be harmful and even  fatal

o Beneficial mutations = organisms can quickly adapt, survive  and reproduce (these mutations cause change in populations) ▪ Any change that is advantageous during selection  We also discuss several other topics like What does zimbalist mean?

pressure will be kept by the population

• Genetic Recombination and Transfer

o Genetic Recombination: when an organism gets & expresses  genes from another organism, is an exchange of nucleotide  sequences between 2 DNA molecules, usually involves  

segments that are identical or nearly identical  

▪ Three forms of Genetic Recombination in Bacteria:

???? Conjugation: transfer of a plasmid from donor cell  

(gram-negative cell donor is a cell with a fertility  

plasmid/F+ cell) to recipient cell (cell without a  

fertility plasmid/F- cell) with a direct connection

(pilus – tube where DNA passes through). Once  the F- cell gets the plasmid from the F+ cell it  becomes a F+ cell too.  

• In cells where the plasmid transferred  doesn’t stay in the cytosol but instead is  incorporated into the cellular chromosome  = High Frequency Recombination Cells (HFR  cells)

o In these cells when they perform  

conjugation a portion of the  

chromosome and a portion of the  

fertility plasmid are transferred to the  

recipient.  

???? Transformation: organism takes up DNA from its  environment, DNA fragments from lysed cells are  accepted by the recipient cell and the genetic  code is acquired by the recipient

• Donor and recipient cells can be unrelated,  is a useful tool in recombinant DNA  

technology  

???? Transduction: when a bacteriophage injects its  DNA into the host cell & degrades the host cells  DNA, when creating new phages if one takes up  part of the host cells DNA instead of its own it is  called a transducing phage and when it injects its  DNA into a new cell the old host’s DNA is  incorporated into the new cells DNA.

• Two Types:

o Generalized Transduction: random  

fragments of host DNA are picked up  

during phage assembly, any gene can  

be transmitted this way  

o Specialized Transduction: A specific  

piece of the host genome is  

incorporated into the virus

▪ Here the phage goes into  

lysogeny & is incorporated into  

the bacteria cell’s genome,  

something happens to make it  

reenter the lytic cycle. The  

excised phage DNA contains  

some host DNA and when new  

phages are produced it has  

some bacterial DNA. All phages  

can function as phages and  

when a new recipient cell is  

infected the bacterial DNA is  

transferred. The DNA (with  

bacterial DNA and phage DNA)  

can enter and do lysogeny or  

just the bacterial DNA can be  

incorporated into the cells  

genome.  

o Transposition: Mutation in which a genetic segment is  

transferred from one location in the genome to another via a  Transposon. This is transfer of genetic information within one  cell.

▪ Transposons are the segments of DNA that are able to  move from one location to another in the genome.  

“Jumping Genes”  

???? Causes rearrangement of the genetic material  

(like a frameshift mutation)  

???? Can occur between plasmids and chromosomes  

and within and among chromosomes  

???? Can be beneficial or harmful, is a way to get  

variation in genomes

???? They go to another location but also stay in their  

original location  

Controlling Microorganisms

• Targets are microorganisms that can infect or cause spoilage. Ex:

o Vegetative bacterial cells & endospores

o Fungal hyphae and spores, yeast  

o Protozoan trophozoites & cysts

o Worms

o Viruses

o Prions

• Ranking in terms of Most Resistant to Least Resistant to  chemical controls  

o Prions – part of microbes, pathogenic proteins that are  tolerant to steam

o Bacterial endospores

o Mycobacteria

o Cysts of Protozoa

o Active-stage protozoa (trophozoites)

o Most Gram-negative bacteria – have outer membrane as well  as the peptidoglycan layer  

o Fungi

o Non-enveloped viruses

o Most gram-positive bacteria

o Enveloped viruses – capsid is not as rigid because of  membrane, if it were more tightly packed it would be more  resistant  

• Terminology and Methods of Control

o Sterilization: process that destroys ALL viable microbes, this  includes viruses and endospores

o Disinfection: process that destroys vegetative pathogens but  not endospores (this is used on inanimate objects, not the  body)  

o Antiseptic: disinfectants that are applied directly to the body  o Sanitization: cleansing techniques that mechanically remove  microbes, this destroys some things  

o Degermation: use mechanical means to reduce the number of  microbes, example would be hand washing with soap and  water

• Microbial Death – how do we tell a living microbe from a dead one?

o Difficult to detect this because microbes don’t reveal vital  signs to begin with, can tell w/membrane functions  sometimes

o One way to tell a microbe is dead is if it has permanent loss  of reproductive capability, even when in optimum growth  conditions  

o Suffixes:

▪ -cide = the death of something  

▪ -static = bacteria are not growing and are also not  dying, there is no change.  

o Factors that Affect Death Rate (how effective an agent is,  is determined by several factors)

▪ Number of Microbes

▪ Nature of Microbes in the population – ex. spores are  harder to kill, enveloped viruses are easier to kill than  naked  

▪ Temperature and pH of the environment - warm  disinfectants work better than cool ones (chemicals  react faster at higher temperatures), acidic conditions  enhance the antimicrobial effect of heat, some  

household disinfectant are more effective at a lower pH  ???? This affects the denaturing of proteins

▪ Concentration or dosage of the agent

▪ Mode of action of the agent – cellular targets of physical  and chemical agents  

???? The Cell Wall: wall becomes fragile and cell lyses,  done by some antimicrobial drugs, detergents, &

alcohol.  

???? The Cell Membrane: membrane loses integrity by  influencing the stability of the phospholipid  

bilayer, detergent surfactants do this.  

???? Protein and Nucleic Acid Synthesis: prevent  

replication, transcription, translation, formation of  peptide bonds, and protein synthesis. This is done  by chloramphenicol, ultraviolet radiation, and  

formaldehyde.

???? Disruption or Denaturing of Proteins: Done by  alcohols, phenols, acids, and heat.  

▪ Presence of solvents, organic matter, or inhibitors o Decimal Reduction: the time it takes to kill 90% of a  population, if start with a big number even a 90% reduction  would still leave a significant amount.  

o Methods of Physical Control  

▪ Heat: denatures proteins, interferes with the integrity o  of the membrane and walls, disrupts of the function and  structure of Nucleic Acids.  

???? Moist: Commonly used to disinfect, sanitize,  

sterilize, pasteurize, kills cells by denaturing  

proteins & destroying cytoplasmic membranes.  

More effective than dry heat because water  

conducts heat better than air.

• Boiling: kills vegetative cells of bacteria &  

fungi, trophozoites of protozoa & most  

viruses within 10 min.  

o Boil at 100°C for 30 minutes to  

destroy non-spore forming pathogens  

• Pasteurization: use heat to kill pathogens &  

reduce number of spoilage microorganisms  

in food & drinks  

o A way to control bacterial population  

so yeast is the dominating population  

without damaging the grapes

o 63°-66°C for 30min – Batch method

that kills a lot of microbes (old  

method)

o 71.6°C for 15 seconds – Flash Method

(New method, controls a lot of  

microbes)

o Not Sterilization – shelf stable milk – 

hit with high temperatures and 

changes flavor

• Autoclaving (like a pressure cooker):

sterilize chemicals & things that can tolerate  moist heat, prevents the escape of steam increasing the pressure, applying pressure  sterilizes because the temperature that  

water boils at increases as pressure  

increases. Destroys all microorganisms in  15 min.  

???? Dry: used for powders and oils, things that can’t  be sterilized with steam or boiling, Denatures  proteins, fosters oxidation of metabolic & structural chemicals. Requires higher temps for  longer times than moist heat because dry heat penetrates slower.  

• Dry Oven: takes longer than an autoclave  (15 min. vs. 16 hours), 150°-180°C  

coagulates proteins

• Incineration: fosters oxidation, alters  protein structure, ultimate means of  

sterilization, ex. Heating a loop in a Bunsen  burner

???? Thermal Death Measurements

• Thermal Death Time (TDT): shortest length  of time to kill all microbes at a certain  

temperature

• Thermal Death Point (TDP): lowest  

temperature to sterilize something within 10  minutes

• Thermal Death Times of Endospores  

o Moist Heat (121°C):  

▪ Bacillus subtilis – 1 min

▪ Clostridium botulinum – 10 min

o Dry Heat (121°C and 120°C)

▪ Bacillus subtilis – 120 min

▪ Clostridium botulinum – 120  

min

▪ Cold Temperatures: slows microbial growth ???? Refrigeration 0-15°C and freezing below 0°C – preserves microbes, used to preserve food, media  and cultures

???? Microbiostatic: slows the growth of microbes ▪ Desiccation: a way to preserve microbes, a gradual  removal of water from cells which leads to metabolic  inhibition – the cells can’t do anything but they are not  dead, preserve them by freeze drying,  

???? Can mail them this way & they don’t grow while  like this.  

???? Not an effective method of microbial control  because they can grow again once water is  

reintroduced.  

???? Lyophilization: freeze drying; preservation  ▪ Radiation: release of high-speed subatomic particles or  waves of electromagnetic energy from atoms ???? Ionizing: electromagnetic radiation  

w/wavelengths shorter than 1nm are ionizing  because when they hit molecule have enough  energy to eject electrons from atoms create ions  • A form of cold sterilization

• This disrupts h-bonding, oxidizes double  

covalent bonds, create highly reactive  

hydroxyl radicals.

• Ions denature other molecules and cause  fatal mutations & cell death.

• Ex: cathode rays, gamma rays, some x-rays • Used to sterilize medical supplies and food  products that can’t take heat (like plastics)  

???? Non-ionizing: electromagnetic radiation  

w/wavelength greater than 1nm, not enough  energy to force electrons out of orbit, excites  electrons to force creation of new covalent bonds  which affects structure of proteins and nucleic  acids.

• Ex: UV light, visible light, infrared radiation,  

radio waves  

o UV light used in microbial control by  

making thymine dimers that prevent  

replication of genetic material & cause  

cells to die. Does not penetrate well,  

disinfects air, transparent fluids, &  

surfaces

▪ Filtration: passage of fluid through a sieve that traps  particles (cells or viruses) and separates them from the  fluid.  

???? sterilize heat sensitive materials & estimate the  number of microbes in a fluid (count how many  

on filter after pouring certain amount through)  

???? HEPA filters used in labs, filter the air in the hoods  so the air in the hood is sterile and the exhaust of  the hood is filtered too

• Pressure system is used by these to control  

the air flow and control what leaves

o Chemical Microbial Control

▪ Types of Chemicals  

???? Disinfectants: do not guarantee all pathogens are  eliminated, used on inanimate objects, don’t kill  

endospores or some viruses, more concentrated  

than antiseptics and can be left on surfaces for  

longer  

???? Antiseptics: chemical used on skin or other tissue ???? Sterilants: chemicals that sterilize, denature  

proteins and DNA by cross-linking organic  

functional groups

???? Degermers: remove microbes but don’t kill them  ???? Preservatives

▪ Desirable qualities of chemicals:

???? Rapid action at a low concentration

???? Toxic to other things, not you (low toxicity)

???? Doesn’t stain, not corrosive

???? Affordable & readily available

???? Water soluble/Alcohol soluble

???? Stable  

▪ Levels of Chemical Decontamination

???? High-level Germicides: kills endospores, use on  something that needs to be sterile but can’t be  heated

???? Intermediate-level: Doesn’t kill endospores, will  kill fungal spores (tuberculosis, virus), disinfects  not sterilize, good for things that come into  

contact with mucous membrane  

???? Low-level: will only kill vegetative cells & normal  fungal cells not spores, ok if just touching but not  used for things that touch mucous membrane &  are invasive

▪ Factors that Affect Germicidal Activity of Chemicals  ???? Nature of material being treated – some things  can’t handle certain chemicals  

???? Degree of contamination – where things are going  is important, are they going to be in contact with  the mucous membrane etc.

???? Time of Exposure – Ex: Alcohol sanitizer needs to  be in contact with your hands for 10 seconds,  time microbes are exposed to chemicals affects  how effective the chemical is.  

???? Strength and Chemical Action of the Germicide  ▪ Examples of Chemicals

???? Halogens  

• Chlorine: Cl2, hypochlorites (Chlorine  

Bleach), chloramines (safer than chlorine),  

one of the most highly used  

o Method: denatures proteins by  

disrupting disulfide bonds  

o Intermediate Level

o Unstable in sunlight, inactivated by  

organic matter – if something is super

dirty it might not be as effective  

against it

o Use: decontaminate water, sewage,

wastewater, inanimate objects

• Iodine: I2, iodophors (betadine)

o Method: interferes with disulfide  

bonds of proteins

o Intermediate Level  

o Use: milder medical and dental  

degerming agents, disinfectants,  

ointments  

o *Staining is a downside

???? Phenolics

• Method: disrupts cell walls & membranes,  precipitates proteins  

o At high concentrations – disrupts cell  walls and proteins

o At low concentrations – disrupts  

critical enzyme systems  

• Low to intermediate-level – Bactericidal,  fungicidal, virucidal, not sporicidal (Strong antimicrobial but not sporocidal)  

• Because of high toxicity don’t use them as  antiseptics anymore

• Ex: Lysol, Triclosan – antibacterial additive  to soaps  

• Use: in lab use phenols to remove proteins  from preps  

• Lister used carboxylic acid (a types of  phenol) to clean skin but was really harsh  skin

???? Chlorohexidine – used instead of phenols, low  toxicity

• Method: surfactant & protein denaturant  w/broad microbicidal properties

• Low to intermediate level – good against  bacteria, variable against viruses & fungi,  Not good on endospores

• Use: skin degerming agents for  

preoperative scrubs, skin cleaning & burns  • Ex: Hibiclens & Hibitane (used to clean skin)  ???? Alcohols

• Method: acts as surfactants dissolving  membrane lipids & coagulating proteins of  vegetative bacterial cells & fungi  

• Intermediate Level – No impact on  

endospores, does impact fungal spores

• Ex: Ethyl, isopropyl in solutions of 50-95% - 70% is more effective than 100% because  100% is dehydrating.  

???? Hydrogen Peroxide

• Method: Produces highly reactive hydroxyl free radicals which damage protein & DNA  while decomposing to oxygen gas – toxic to  anaerobes

• At low concentrations: Antiseptic

• At high concentrations: Sporicidal  

???? Aldehydes

• Method: Glutaraldehyde & formaldehyde kill  by alkylating protein & DNA  

• High level – Glutaraldehyde in 2% solution  (Cidex) is used as a sterilant for heat  

sensitive instruments  

o 2nd chemical to use as a sterilant

• Intermediate level – Formaldehyde is a  disinfectant & preservative, limited use due  to toxicity, works fast

o Formalin – 37% aqueous solution  

???? Gases and Aerosols

• Method: strong alkylating agents

• High Level

• Use: sterilize & disinfect plastics &  

prepackaged devices, foods  

o Used in a chamber w/temperature,  

pressure & oxygen controlled  

o 19-3 hours are needed to aerate,

sterile air for hours after  

o gas is explosive & can harm our  

bodies & cause cancer  

• Ex: ethylene oxide and propylene oxide  ???? Detergents and Soaps  

• Method: quaternary ammonia compounds  (quats) work as surfactants (polar  

molecules that have hydrophobic and  

hydrophilic parts), change some fungi &  bacteria membrane permeability

• Very low level  

• Use: soaps are a method of degerming,  mechanically remove soil & grease that  contains microbes  

o Good against viruses = Anti-viral

o Most common in Bactene

o Non-germicidal soaps – cause  

inhabitant microbes to be pulled out  

of skin and after days of scrub water  

sitting the number of microbes are  

higher with these soaps (which is the  

category that antibacterial soaps are  

under) than with Germicidal Soap

o Germicidal Soaps – with prolonged  

used the number of bacteria  

decreases.  

???? Heavy Metals

• Method: low concentrations of silver and  mercury solutions kill vegetative cells by  inactivating proteins

o Oligodynamic action: bind to the final  

groups of proteins to inactivate them  

o Have antimicrobial properties

• Low level  

• Ex: Merthiolate, silver nitrate, silver  

???? Dyes as Antimicrobial Agents

• Method: aniline dye very active against  

gram-positive species of bacteria and  

various fungi  

• Low level, narrow spectrum of activity

• Use: antisepsis & treat wounds

???? Acids & Alkalis

• Low level

• Organic Acids: prevent spore germination,  

bacterial growth, fungal growth

• Acetic Acid: inhibits bacterial growth

• Propionic Acid: slows molds – this is the  

thing that propionic bacteria makes that  

gives cheese its flavor  

• Lactic Acid: prevents anaerobic bacterial  

growth – lactobacillus ferments glycogen  

and decreases pH = keeps the vagina acidic

• Benzoic Acid: inhibit yeast

Antibiotics  

• Principles of Antimicrobial Therapy  

o Give a drug to an infected person, it destroys the infective  agent without harming the host’s cells  

▪ Therapeutic Index is good when antimicrobial has an  

increased toxicity to microbes and decreased toxicity to  

us/humans

???? Therapeutic index: the ratio of the dose of the  

drug that is toxic to humans as compared to its  

minimum effective dose (Ratio= smallest effective  

dose/amount that is toxic to humans) ???? want #  

on bottom to be smaller  

• Higher index is desirable

o Antimicrobial drugs are produced naturally or synthetically  ▪ *Side Note: The mortality rate in some countries is the same as before antibiotics because of high rates of  

infant mortality due to lack of access to healthcare  

• Characteristics of an Ideal Antimicrobial  

o Selectively Toxic

o Microbicidal – kills microbes

o Relatively soluble – functions when highly dilute, makes sense  because we are made of mostly water

o Remains potent long enough – not broken down or excreted  prematurely, with penicillin this was a problem early on  because people were peeing it out  

o Doesn’t lead to resistance – this is difficult

o Complement or assists the activities of the host defenses o Active in tissues and body fluids  

o Can get to site of infection

o Affordable  

o Doesn’t negatively affect host’s health…allergies or other  infections

• Domagk – showed that red dye called prontosil could be active  against bacteria – this was the first sulfur drug

• Paul Erlich – some dyes dye the microbes and not the tissue, he  came up with an early chemotherapy for syphilis

• Terminology

o Prophylaxis: Action taken to prevent disease, with a specified  means against a specified disease

o Chemotherapeutic drug: drugs that act against diseases o Antimicrobial: any compound used to treat infectious disease,  may also function as intermediate-level disinfectant  

o Antibiotic: antimicrobial chemicals produced naturally by  microorganisms  

o Synthetic/Semi-Synthetic: Antimicrobials that are completely  synthesized in the lab

o Narrow Spectrum: Drugs that work against only a few kinds  of pathogens, target a specific cell component that is only  found in certain microbes  

o Broad Spectrum: Drugs that work against many different  kinds of pathogens, target cell parts that are common to most  pathogens (ribosomes)  

o Spectrum: range of activity of a drug  

• For antibiotics to work must work against the differences in the cells  o For example antimicrobial or antibiotics works against the  peptidoglycan in bacteria

• Antibiotics come from one genus, are natural in origin – we are not  very good at creating drugs on our own  

• Sporulation (Life cycle of streptomycin)

o Exospore ???? Spore Germination ???? Vegetative Hyphae Growth  (digs into the plate) ???? Aerial Hyphae Growth (microbe grows  up and out of the media, produces antibiotic as it grows aerial  hyphae, when it does this it breaks down substrate hyphae  for nutrients and the nutrients go into the soil) ???? Septation  (growth subdivides and twists) ???? Spores maturation

• Theory for why organisms make Antibiotics  

o Competitive advantage idea

o Sub-inhibitory levels of antimicrobials cause response to  things around it  

o Junk Mechanism  

o Evolutionary leftover mechanism ???? this is the reason we have  resistance ???? prokaryotes are the main producers of  

antibiotics, they can kill other things in the same genus, these  microbes have a mechanism of resistance ???? can’t get rid of  the resistance  

• Interactions Between Drug & Microbe

o Antimicrobial drugs should be selectively toxic – drugs should  kill or inhibit microbial cells without damaging the host tissue  ▪ When the characteristics of the infectious agent become  similar to the host cell selective toxicity becomes  

difficult to achieve = more side effects  

• Mechanisms of Drug Action

o Inhibition of Cell Wall Synthesis

▪ Β-lactams  

???? Basic Structure: beta-lactam rings

???? Mode of Action: inhibit peptidoglycan formation by  binding to the enzymes that cross-link NAM  

subunits (attacks between the NAG/NAM  

subunits). This causes the bacterial cells to have  

weakened cell walls as they grow and they are  

not resistant to osmotic pressure, as water moves  into the cell, the membrane bulges through the  

weakened part of the cell wall & the cell  

eventually lyses.  

• For something that attacks to the forming  

bonds we need something to be growing  

rapidly for this to work (aka cause cells to  

lyse)  

???? Effectiveness:

• Penicillinase or B-lactamase

???? Drugs within this group:

• Penicillins – Penicillin chrysogenum is a  

major source  

o Consists of three parts:

▪ Thiazolidine Ring

▪ Beta-lactam ring

▪ Variable side chain dictating  

microbial activity – affect the  

ability to get across the outer  

membrane  

o Subgroups and Uses of Penicillins

▪ Penicillin G and V most  

important natural forms  

▪ Drug of choice for Gram-Positive  

cocci (streptococci), some  

Gram-Negative bacteria  

(meningococci and syphilis  

spirochete)

▪ Semisynthetic Penicillins –

ampicillin, carbenicillin,  

amoxicillin broader spectra –

Gram-Negative infections  

▪ Penicillinase-resistant –

methicillin, nafcillin, cloxacillin  

(resistant to enzyme that  

breaks down penicillin)

▪ Primary problems: allergies to  

penicillin and resistant strains of  

bacteria  

• Cephalosporins: Targets the building of  peptidoglycan, 1/3 of all antibiotics  

administered

o Relatively broad spectrum, resistant  to most penicillinases, cause fewer  

allergic reactions  

o Some given orally, many parentally  o Generic Names have root – cef, ceph,  kef

o 4 Generations Exist (each group more  effective against gram-negatives than  the previous one, better dosing  

schedule and less side effects) –

become more broad range as  

progress:

▪ First Generation: cephalothin,  

cefazolin – most effective  

against gram-positive cocci and  

few gram-negative  

▪ Second Generation: cefaclor,  

cefonacid – more effective  

against gram-negative  

▪ Third Generation: cephalexin,  

ceftriaxone – broad-spectrum

activity against enteric bacteria  

w/beta-lactamases

▪ Fourth Generation: cefepime,  

widest range, both gram

negative & gram-positive  

• Carbapenems: Imipenem – broad spectrum  drug, used for infections w/aerobic &  

anaerobic pathogens, low dose,  

administered orally, few side effects  

• Monobactams: Aztreonam – narrow  

spectrum drug, used for infections by gram

negative aerobic bacilli, used by people  

allergic to penicillin

▪ Non beta-lactam Cell Wall Inhibitors  

???? Vancomycin: Narrow-spectrum, used for  

staphylococcus infections when there is resistance  to penicillin & methicillin or if patient is allergic to  penicillin, is toxic and hard to administer, use is  restricted

• Disrupts formation of G-positive cell wall by  interfering with alanine-alanine crossbridges  linking NAG subunits

• Used to treat MRSA

• Now some things are resistant to this  

???? Bacitracin: narrow-spectrum, made by strain of  Bacillus subtilis, used in topical ointments  

• Blocks NAG/NAM secretion from the  

cytoplasm

• Main ingredient in neosporin

???? Isoniazid (INH): interferes w/mycolic acid  synthesis, treats infections w/Mycobacterium  tuberculosis

• Disrupts the formation of arabinogalactan mycolic acid by mycobacteria  

• Used in a triple therapy to treat  

Tuberculosis

o Breakdown of cell membrane structure or function: cell dies  from disruption in metabolism or lysis  

▪ Cant carry out chemical rxns without intact membrane ▪ These drugs have Specificity for particular microbial  group based on differences in types of lipids in  

membranes  

▪ Polymyxins: interact w/phospholipids, cause leakage,  specifically in gram-negative bacteria

▪ Amphotericin B and Nystatin: make complexes with  sterols (ergosterol) on fungal membranes ???? disrupts  the membrane and causes lysis = leakages (fungicidal)  

???? The difference between fungal cells and your cells  are the sterols, membranes without cell walls  

have sterols to maintain rigidity.  

???? Human cells somewhat susceptible because  

cholesterol is similar to ergosterol (but don’t bind  as well)

o Inhibition of nucleic acid synthesis, structure or function ▪ Block synthesis of nucleotides, inhibits replication, stops  transcription  

▪ Chloroquine: binds and cross-links the double helix,  inhibit DNA helicases  

▪ Antiviral drugs, analogs of purines and pyrimidines  insert in viral nucleic acids, prevents replication  

▪ Rifampin: bind and inhibit the action of RNA polymerase  during synthesis of RNA from DNA, binds more readily  to prokaryotic than eukaryotic so more toxic to those.  o Drugs that Act on DNA or RNA  

▪ DNA Gyrase Inhibitors  

???? Fluoroquinolones: synthetic drugs that are active  against bacterial DNA, work by binding to DNA  

gyrase (enzyme necessary for correct coiling and  uncoiling of replicating bacterial DNA) and  

topoisomerase IV, may act against replication of  

mitochondrial DNA in some Eukaryotes.  

• Broad spectrum effectiveness

• Concerns w/overuse of quinolone drugs –

recommend careful monitoring to avoid  

ciprofloxacin-resistant bacteria

o Inhibiting Protein Synthesis – Ribosomes of euk. differ from  prok. Antimicrobics selective action against prokaryotes but  can also damage the euk mitochondria

▪ Aminoglycosides: ex. Streptomycin (treats TB),  gentamycin. These insert on sites on the 30S subunit,  cause misreading of mRNA  

▪ Tetracyclines: 4 ring structure, block attachment of  tRNA on A acceptor site & stop further synthesis, used  against lime disease, Typhus, some STD’s, has side  effects

▪ Chloramphenicol: phenol ring in structure, affects  peptide bonding, attaches to 50S subunit and prevents  peptide bond formation  

???? Broad spectrum  

???? Nitrobenzene type structure

???? Harms bone marrow

???? Used for rickettsia and chlamydia  

▪ Macrolides – Erythromycin: binds to 50S subunit  blocking proper movement of mRNA through ribosome,  synthesis stops  

???? Used prophylactically – before someone has an  infection

???? Used for penicillin resistant organisms

o Blocks key metabolic pathways  

▪ Sulfonamides & Trimethoprim block enzymes required  for tetrahydrofolate synthesis that is needed for DNA  and RNA synthesis  

???? Methods

• Competitive inhibition: the drug competes  

with normal substrate for the enzyme’s  

active site

• Synergistic Effect: drugs work better  

together than on their own

▪ *Remember that the problem with broad spectrum  antibiotics is that they kill normal microbiota too, which  can be good/helpful to us

• Agents to Treat Fungal Infections  

o Fungal cells eukaryotic, drug toxic to fungal cells also toxic to  human cells = hard to treat, bad side effects, remember  antibiotics don’t work on fungal organisms  

o Five Antifungal Groups:

▪ Macrolide polyene  

???? Amphotericin B: mimic lipids, most versatile &  

effective, topical and systemic treatments  

???? Nystatin: topical treatment and oral swishes

• Again the function of these make complexes  

w/sterols (ergosterol) on fungal membranes  

???? disrupts the membrane & causes lysis =  

leakages (fungicidal)  

▪ Griseofulvin: used for stubborn cases of dermatophyte  infections, is nephrotoxic (damaging to the kidneys)

▪ Synthetic azoles: broad spectrum, ketoconazole,  

clotrimazole, miconazole  

▪ Flucytosine: analog of cytosine, used against cutaneous  mycoses (disease of the hair, skin, and nails) or used  with Amphotericin B for systemic mycoses  

▪ Echinocandins: damages cell walls, used against  

capsofungin, these are the reason they stopped  

antifungal research because they are very good against  dominant fungal infections.

• Antiparasitic Chemotherapy

o Antimalarial drugs: quinine, chloroquinine, primaquine,  mefloquine

o Antiprotozoan drugs: metronidazole (Flagyl), quinicrine,  sulfonamides, tetracyclines

o Antihelminthic drugs: these immobilize, disintegrate, inhibit  metabolism

▪ Mebendazole, thiabendazole –broad spectrum- inhibit  function of microtubules, interfere w/the usage of  

glucose & disables them

▪ Pyrantel, piperazine – paralyze muscles  

▪ Niclosamide – destroys scolex

• Antiviral Chemotherapy

o Selective toxicity hard because of obligate intracellular  parasitic nature of viruses

o Methods

▪ Inhibition of virus entry or release (interfere with fusion  of virus to the membrane)

???? Fuzeon: blocks HIV infection

???? Amantidine: blocks influenza virus

???? Tamiflu and Relenza: stops actions of influenza  

neuramidase required to enter the cell  

▪ Block replication, transcription, or translation of viral  genetic material – Inhibition of nucleic acid synthesis

???? Nucleotide analogs  

• Acyclovir – used against herpes virus

(terminates DNA replication)

• Ribavirin – a guanine analog used against  

RSV, hemorrhagic fevers  

• AZT – thymine analog used against HIV

(HIV is an RNA based virus, if block reverse  

transcriptase, which converts RNA to DNA,  

then can’t produce viral DNA to enter the  

host DNA)  

▪ Prevent maturation of the viral particles – Inhibition of  Effective Viral Assembly and Release  

???? Protease inhibitors – used against HIV (inserts  

into HIV protease an enzyme that clips viral  

proteins into functional pieces)  

• Interferons (INF) – human-based glycoproteins, made mostly by  fibroblasts and leukocytes  

o Therapeutic benefits:

▪ Decrease healing time and complications of infections – antiviral & anticancer properties

▪ Prevents/reduces symptoms of cold & papilloma virus  

▪ Slows progress of some cancers, leukemia, &  

lymphomas

▪ Treats hepatitis C, genital warts, Kaposi’s sarcoma

Drug Resistance – an adaptive response when microbes begin to tolerate  an amount of drug that would normally kill them, because of genetic  versatility or variation (can be intrinsic or acquired)

• Acquired Resistance

o spontaneous mutations in critical chromosomes

o get new genes/sets of genes from resistance factors (R  plasmids) encoded w/drug resistance, transposons

▪ Antibiotic Resistance Transfer – based on R plasmids  

(plasmids carrying resistance genes)  

• Mechanisms of Drug Resistance  

o Mechanism 1: limiting access of the antibiotic due to  

decreased permeability to drug/increased elimination of drug  from cell – acquired/mutation

▪ Outer membrane porins 

▪ Active Efflux: resistant cells can pump the antimicrobial  out of the cell before the drug can act. Some cells are  

able to pump more than one antimicrobial from the cell.  

▪ Reduced uptake across cytoplasmic membrane  

o Mechanism 2: Drug Inactivation due to acquired enzymatic  activity – penicillinases – acquired mutation

▪ B-lactamases: enzymes break the beta-lactam rings of  penicillin & similar molecules making them inactive

???? In Penicillin the efficacy of it as an antibiotic is  

dependent on the Lactam Ring and penicillinases  

breaks that ring to make it inactive  

▪ Modifying Enzymes  

o Mechanism 3: Modification or protection of target -

acquired/mutation

▪ Resistant cells may alter the target of the drug so that  the drug either cannot bind to it or binds less effectively ???? change in drug receptors

o Mechanism 4: Change in metabolic Patterns – mutation of  enzyme  

▪ Alter the metabolic chemistry or abandon sensitive  metabolic step altogether, cell could become more  

resistant to a drug by producing more enzyme  

molecules for the metabolic pathway & reducing the  

power of the drug

▪ Trymethoprin and Sulfonamides

• Development of Resistance  

o Spontaneous Mutation

o Transfer of Resistance – 3 Methods

▪ Transduction: when a bacteriophage injects its DNA into  the host cell and degrades the host cells DNA, when  

creating new phages if one takes up part of the host  

cells DNA instead of its own it is called a transducing  

phage and when it injects its DNA into a new cell the  

old host’s DNA is incorporated into the new cells DNA.

▪ Transformation: organism takes up DNA from its  

environment, DNA fragments from lysed cells are  

accepted by the recipient cell and the genetic code is  

acquired by the recipient

▪ Conjugation: transfer of a plasmid from donor cell  

(gram-negative cell donor is a cell with a fertility  

plasmid/F+ cell) to recipient cell (cell without a fertility  plasmid/F- cell) with a direct connection (pilus – tube  where DNA passes through).

• Factors that Contribute to Resistance  

o Natural Selection

▪ Big populations of microbes are likely to have drug  resistant cells from prior mutations or transfer of  

plasmids, there is no growth advantage to these until  they are exposed to a drug.

▪ Once exposed sensitive cells die & resistant cells survive

▪ Population becomes resistant from selective  

pressure/natural selection

▪ Indiscriminate use of antimicrobials worldwide has led  to many drug resistant microorganisms.  

o Health care mentality

o Agriculture

o Worldwide Resistance

• How to Limit Drug Resistance

o Drug Usage

▪ Physicians – accurate diagnosis, give right drug

▪ Patients – comply w/guidelines

▪ Combined Therapy – more than 1 antibiotic works to kill  off microbes, remember dead cells can still give other  cells nucleic acids

o Drug Research

▪ Develop shorter duration, higher dose antimicrobials ▪ Find drugs whose structures are not inactivated by  enzymes & not readily circumvented

o Long-term  

▪ Educate Healthcare workers – reduce the abuse of  antibiotics

▪ Restrict use of antibiotics

▪ Stop using antibiotics in animal feed

▪ Come up w/programs that have effective therapy  

available to low income populations

▪ Vaccine where possible – there is nothing out there  supporting a link between autism & vaccines

???? Some components of vaccines work better with  

others, they are synergistic so vaccines work  

better if taken together

• Interactions Between Drug & Host

o ~5% of everyone taking antimicrobials will experience serious  adverse reaction/side effects  

o Major Side Effects:

▪ Damage to tissue because of drug toxicity

???? If use Tetracycline when child is developing =  

tooth discoloration

???? Flagyl - hairy tongue is a side effect of this  

antibiotic because it kills good microbes that were  

stopping that before

▪ Allergic Reactions

▪ Disrupts balance of normal flora = superinfections  

possible (Secondary infections that are caused by  

treatment of antibiotics which are killing microbes and  opening niches)  

• What to Consider when Selecting a Drug

o Identify microorganism causing infection – restrict the use of  anything that might work against an ideal organism

▪ Identify as soon as possible

▪ Specimens should be taken before starting  

antimicrobials  

o Test microorganism’s susceptibility to drugs in vitro when  indicated

▪ Essential for bacteria that are commonly resistant  

▪ Kirby-bauer disk test: create lawn of microbe and  

inoculate with a disk of antibiotic, read the zone of  

inhibition to determine if the microbe is sensitive or  

resistant

???? Resistant: If something is resistant then we can’t  

get the antibiotic to a high enough concentration  

in the patients to safely kill the microbes

???? Intermediate: Can use in patients but will have  

side effects

???? Sensitive: a low enough concentration of the  

antibiotic kills the microbe that it could be used in  

a patient

▪ E-test diffusion test

▪ Dilution Test: conducted to find the MIC (minimum  inhibitory concentration) – the smallest concentration of  a drug that will visibly inhibit the microbes growth  

▪ Provide profile of drug sensitivity

o Overall condition of the patient – will drugs have a side effect  • MIC and Therapeutic Index  

o MIC: Minimum Inhibitory Concentration = the smallest  concentration of a drug that will visibly inhibit the microbe’s  growth  

o The in vitro activity of a drug is not always correlated to in  vitro effect

▪ If therapy fails consider a different drug, combination of  drugs or different administration of drugs  

o It is best to choose a drug with highest level of selectivity and  lowest level of toxicity (determined by Therapeutic Index)

▪ Therapeutic Index: the ratio of the dose of the drug  

that is toxic to humans as compared to its minimum  

effective dose (Ratio= smallest effective dose/amount  

that is toxic to humans) ???? want # on bottom to be  

smaller  

???? Higher index is desirable

• Final Points on Antibiotic Resistance  

o Antibiotics still valid therapy  

o Education is vital – patients can’t demand drugs, need to take  their full prescription

o Combinations of drugs & developing new vaccines = slows spread of drug resistant pathogens  

Relationships between Microbes and their Hosts

*Human body always in state of dynamic equilibrium, many interactions  between human and microbe involve biofilms (on teeth, tongue, periodontal  pockets), colonization of the body by microbes is a constant give and take.  • Contact, Colonization, Infection, Disease

o Normal (resident) Flora/Microbiota: the normal colonizing  bacteria or microbes

▪ Areas in contact w/outside environment typically harbor  resident microbes  

▪ Internal organs, tissues, & fluids microbe free

▪ Transients: Microbes that occupy body for short periods  of time (temporary, a lot of infectious diseases are  

transient or temporary)  

▪ Residents: microbes that become established

???? Microbial Antagonism: Bacterial Flora benefit the  

host by preventing overgrowth of harmful  

microbes – simple niche occupancy

???? Endogenous Infections: infections from normal  

flora being introduced to a site that was sterile –

there is resistance because you are already  

colonized  

▪ *Remember how different all the environments of our  body are, the norm is that certain microbes live in  

specific conditions, it is rare that a microbe lives in  

many different conditions in the body.

o Infection: something coming from outside or conditions of  being pathogenic

o Pathogen: something that has pathogenic ability

o Infectious: capable of causing infection  

• Initial Colonization of the Newborn: before birth you are not  colonized, birth is when begin being colonized by mom – being  breastfed vs formula leads to biases of different colonies within the  child  

• Flora of the Human Skin

o Skin is a barrier to infection

o The subcutaneous level is not colonized – this is the boundary  to sterile internal organs  

• Maintenance of the Normal Resident Flora  

o Normal flora essential to health of humans  

o Flora creates an environment that prevents infections &  enhances host defenses – flora can be altered by antibiotic  therapy diet change and disease

o Probiotics: introducing known microbes back into the body  • Infection – Fundamental Events in Pathogenesis

o Portal of Entry: entrance site of pathogenic microorganisms  (skin, mucous membranes, placenta) – Respiratory tract is  the most frequent

o Adherence: process where phagocytes attach to  

microorganisms through binding of complementary chemicals  on cytoplasmic membrane  

o Penetration/Invasion: entrance into the tissue

o Proliferation: rapid increase in numbers/reproduction of a cell,  part or organism

o Pathology: infection of target and disease

o Portal of Exit: exit site of pathogenic microorganisms (nose,  mouth, urethra)

• Types of Pathogen

o True Pathogen (primary pathogens): these make a healthy  individual sick. Ex: influenza virus, plague, bacillus, malarial  protozoan  

o Opportunistic Pathogens: needs an opportunity to infect, like  a cut or weakening of immunity. Ex: Pseudomonas sp &  Candida albicans

o The severity of the disease depends on the virulence (ability  to cause a disease) of the pathogen – Virulence Factors: part  of the microbe that contributes to the ability to cause disease  • Factors that Weaken the Host Defenses

o Age/Extreme Youth

o Genetic defects & acquired defects - mutations in part of the  system makes them more susceptible to the disease  

o Surgery

o Disease – cancer, liver malfunction, diabetes

o Chemotherapy/immunosuppressive drugs  

o Physical/Mental Stress

o Other infections  

• Routes of Entry for Invading Pathogens

o Broken skin or insect bite

o Anus, urethra, vagina or penis

o Placenta

o Mouth, nose, eyes, and ears – mucous membranes

o Some organisms have their own enzymes that facilitate their  entry

• Requirements for an Infectious Dose (ID)  

o Infectious Dose: minimum number of microbes required for  infection to proceed

o Microbes with smaller ID = greater virulence(ability to infect  and cause a disease) - can be as small as one cell, others are  thousands

o If there is no ID then they will not result in infection ▪ If don’t get proper ID then will not have been exposed  and will not get sick  

• Surviving Host Defenses

o Once the pathogen is established it is all about avoiding the  host cells to get to place of infection

o The initial response of the host defenses come from  Phagocytes: cells that take in microbial cells  

o Pathogens use Antiphagocytic Factors to avoid phagocytosis o Leukocidins: things that are toxic to white blood cells and are  produced by species of Staphylococcus and Streptococcus o Slime layer or Capsule: Makes phagocytosis of pathogen by  host cells difficult  

o Some pathogens have the ability to survive being  

phagocytized  

• Exotoxin vs. Endotoxin  

o Exotoxin: Secreted by bacteria to breakdown the host cell, it  has protein targets and a localized effect to specific tissues ▪ Hemolysins

▪ A-B Toxins (A-active, B-binding)

o Endotoxin: lipopolysaccharide (LPS), part of the outer  membrane of gram-negative cell walls. Dead gram-negative  bacteria release endotoxin (lipid A) which induces effects such  as fever, inflammation, diarrhea, shock and blood  

coagulation, not localized, systemic effect  

o Toxicity

▪ Exotoxin: high amount  

▪ Endotoxin: small amount

o Effects on the Body

▪ Exotoxin: Systemic Effect

▪ Endotoxin: Localized Effect

o Chemical Composition

▪ Exotoxin: protein

▪ Endotoxin: component of LPS

o Heat Denaturation

▪ Exotoxin: protein can be denatured

▪ Endotoxin: heat stable

o Toxoid formation:  

▪ Endotoxin: LPS can’t be converted to a toxin  

o Immune Response

▪ Exotoxin: can get an immune response

▪ Endotoxin: no immune response from LPS (lipid A  

portion)

o *All gram(-) have endotoxin and no gram(+) have it.  o *Organisms may or may not have the ability to produce an  enzyme (exotoxin)

• Extracellular Enzymes

o Hyaluronidase and Collagenase  

▪ Invasive bacteria reach the epithelial surface, the  

bacteria produce these enzymes and invade deeper into  the tissues.

???? Breaks down hyaluronic acid & collagen, helps get

through the tissue

o Coagulase and Kinase  

▪ Bacteria produces coagulase, clot forms around the  bacteria so host cells don’t detect it, bacteria later  

produce kinase & dissolve the clot, release the bacteria  ???? Staphylococcus is a good example of something  

that does this

• Stages in the course of infection & disease

o Incubation Period: no signs or symptoms

o Prodromal Period: vague, general symptoms

o Illness: most severe signs & symptoms

o Decline: declining signs and symptoms

o Convalescence: no signs or symptoms  

• Signs and Symptoms of Inflammation

o Signs: measurable, objective evidence of disease noted by an  observer like a doctor

o Symptoms: subjective evidence of disease sensed by the  patient

o Examples of earliest symptoms of disease as a result of  activation of the body defenses are fever, pain, soreness,  swelling

o Signs of Inflammation:  

▪ Edema: accumulation of fluid

▪ Granulomas & abscesses: walled-off collections of  

inflammatory cells & microbes

▪ Lymphadenitis: swollen lymph nodes  

• Signs of Infection in the Blood  

o Change in number of white blood cells  

▪ Leukocytosis: increase white blood cells

▪ Leukopenia: decrease in white blood cells  

o Septicemia: Microorganisms multiplying in blood, present in  large numbers  

o Bacteremia: small numbers of bacteria in blood, not  necessarily multiplying  

o Viremia: small number of viruses, not necessarily multiplying  • Infections that go Unnoticed  

o Asymptomatic: (subclinical) infections that although the host  is infected they don’t show any signs of the disease and don’t  know they are infected (inapparent infection) so they don’t  seek medical attention

• Portals of Exit – pathogens have a specific exit and this exit  influences how the infection spreads

o Normally infectious diseases have a way out but in fungal  infections the infected person has to die and decompose for  the infection to get into the soil & then spread that way.  o Examples of Portals of Exit:

▪ Respiratory: Mucus, sputum, nasal drainage, saliva ▪ Skin scales/flakes

▪ Fecal exit

▪ Urogenital – method for STDs ▪ Removal of blood

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