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Lecture 2

Lecture 2 Continued.docx

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University of Ottawa
Health Sciences
William Yan

Lecture 2 Continued Antibiotic Resistance -the first antibiotic discovered was penicillin -was and continues to be useful but its usefulness has decreased due to antibiotic resistance -saved many lives especially in WWII -conditions were not sterile and wounds were heavily infected by bacterial pathogens -in developed countries, at least 80% of all strains of Staphylococcus aureus are highly resistant to penicillin, even though it has been the choice of treatment -penicillin is no longer used to treat this pathogen due to resistance -long term usage doesn't always lead to resistance (i.e.: S. Pyogenes) -penicillin is more useful against gram-positive bacteria compared to gram-negative bacteria Slide 27: -antibiotics are so useful and so commonly used because they are selective in where and how it acts -antibiotics can usually act on receptors or targets found only on bacteria -no other groups of living organisms have targets that antibiotics can act on -therefore, there are very few side effects -human cells do not have the right targets for antibiotic cells to act on -antibiotics target important cell structures or interfere with key processes that the bacteria use to survive -in order to treat the pathogen, we need to know what organism is causing the infection, where the infection is happening, and if there are any allergies (i.e.: to penicillin) -where and how we administer the antibiotics -i.e.: orally (typically), IV / localized administration to the site of infection if very serious -we need to monitor therapy -it is not enough to prescribe the antibiotics -if symptoms continue, the antibiotics are not working and the strain may be resistant -some antibiotics may cause side effects -some of the most common targets on a bacterial cell are: -cell wall structure (antibiotics break down the cell wall and lyse the bacteria) -protein synthesis machinery (i.e.: enzymes used in protein synthesis, ribosomes) -DNA synthesis machinery (i.e.: enzymes, DNA, etc.) Slide 29: Mechanisms of Resistance 1) Alteration in Drug Target -each antibiotic has a specific site it needs to bind to in order to be efficient -if the site is changed enough so that the antibiotic cannot recognize it or bind to it, it becomes ineffective -this occurs often through genetic mutation -every time a cell divides, there is a chance of DNA mutation (1 in 10 million times) -mutation is random; for mutation to take place right at the DNA location where the target is found, it is very rare -there are so many bacteria and so many multiplications that these events do occur -if there is even 1 resistant E.coli left in the gut, it can thrive because there is no competition -there ends up being many resistant E.coli -sometimes, resistant markers can be transferred from 1 bacteria to another -though E.coli is part of normal flora, if the pathogen picks up the marker from normal flora bacteria, it will then become resistant -selective pressure: applying pressure that selects for the resistant strain that has the advantageous mutation and they can dominate over non-resistant strains 2) Production of Inactivating Enzymes -a bacteria that is normally killed off by antibiotics can become resistant if it picks up a marker that codes for an enzyme that happens to act on antibiotics -some bacteria can make penicillinase which destroys penicillin so it cannot act 3) Decreased Uptake of Antibiotic -taking antibioitics but there is no uptake of antibiotic -due to change in cell wall / permeability / charge -bacteria is impermeable to the antibioitic -bacteria pumps out antibiotic as fast as it can get in *don't need to know table on page 27; know some combinations and general mechanisms* -won't be on MC exam Slide 30: -resistance can be broken down into 2 types: 1) Intrinsic Resistance -there is an inherent characteristic of a bacterium that makes it resistant to a certain antibiotic -they simply do not have the right structure for the antibiotic to act on it -the target site is never there -predictable resistance 2) Acquired Resistance -unpredictable resistance -overtime, bacteria become so resistant that the antibiotic is ineffective *don't need to study page 28 and 29; just read through* Slide 31: -changes in a chromosome in bacterium can lead to multiple resistance -it was thought that you needed antibiotics to apply selective pressure in order to cause resistance -now, exposure to 1 antibiotic can turn on resistance to more than that 1 antibiotic Slide 32: -once a bacterium becomes resistant, what do we do that makes this problem worse? -overuse / misuse of antibiotics cause further resistance -everytime we use antibiotics, we increase chance of mutation -we have to use antibiotics more carefully and only when necessary in order to decrease chance of resistance -a lot of misuse comes from paediatricians who prescribe broad-spectrum antibiotics (this is usually not necessary) and ends up selecting mutant populations of bacteria -e.g.: giving farm animals antibiotics in order to prevent infection is common -by continuously using antibiotics, it is spread into the environment and the number of mutants is increased due to selective pressure; this may eventually come back to human population Slide 33: -we can minimize antibiotic resistance by; -use as specific an antibiotic as possible and only when necessary -do testing first before treatment -don't rely on antibiotics; prevention / education is key Slide 34: -prevention through proper hygiene and handwashing -when we get a prescription for antibiotics, we need to take the prescription for the entire duration instead of stopping after we are feeling better -if we don't finish it, there may be more resistant bacteria taking over -just because symptoms are gone, doesn't mean that all bacteria are gone; bacteria are probably just below the threshold -resistance is not always to the same extent; some bacteria are more resistant than others even if they are in the same population -the gut population is always the first to get hit due to oral administration -bloodstream must carry remaining antibiotics to the site of infection -over time, less and less E.coli remain in the gut so more antibioitic can reach the bloodstream so there is a high concentration of antibiotics in the bloodstream and will reach a maximum potential; this maximum potential will be enough to kill the most resistant bacteria -if the maximum potential is not reached, there are a small number of bacteria that are remaining and these are the highly resistant ones; you made need to switch antibiotics or take a much higher dose to kill of the resistant bacteria -this is a problem especially in developing countries -antibiotics are expensive there -parents try to be efficient with antibiotics; this selects for mutant strains Lecture 3 Diagnostic Microbiology -platinum wire on inoculation loop is used because it heats up and cools down very quickly which is useful when working with culturing bacteria -we need to isolate a pure culture from a specimen -the easiest types of samples to work with are blood samples and CSF which are supposed to be sterile -if there is bacteria there, it is likely that it is causing the symptoms -stool samples are the most difficult to work with because the gut is filled with so much normal flora; the gut also handles lots of food with many different types of bacteria Streaking A Plate for Isolated Colonies -this is a common procedure in microbiology -a petri dish contains a medium (agar) -transfer sample on the surface of the agar -resterilize the loop -retrace original inoculum onto another part of the plate -after many cycles, the number of bacteria should be reduced and at the end, the bacterium should be clones of one another and when transferred to another plate, you should get a pure culture with colonies (which represents 1 million units per colony) Slide 5: Preservation of Cultures -use a refrigerator for short-term storage -for long term storage, you can use liquid nitrogen, freezers, lyophilization -we keep samples long term for research purposes (i.e.: comparing pathogens after many years) Slide 6: Identification -once we have a pure culture we can look at : -colony morphology -we look at the colonies and write down everything we can observe (i.e.: color, size, smell, texture, smooth, bumpy, curved, flat, etc.) -thes
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