Notes for Microbiology Final Exam

Phar­ma­col­ogy refers to drug admin­is­tra­tion (includ­ing dosage) and dis­tri­b­u­tion in the body (phar­ma­co­ki­net­ics). The area under the curve includes T half-life, peak con­cen­tra­tion, time to achieve peak, elim­i­na­tion and pro­tein bind­ing. It’s one of the most impor­tant aspects of drug study, includ­ing antibiotics.

Peak and Trough

Let’s look at the peak and trough of gen­tamycin, which is an amino­gly­co­side. If the con­cen­tra­tion is too high, the peak could lead to oto­tox­i­c­ity and nephro­tox­i­c­ity. So it’s essen­tial you mea­sure peak and trough. If the peaks are exceeded, their mid­dle ear appa­ra­tus are destroyed so they can’t tell their posi­tion and they could have kid­ney failure.

The nor­mal peak and trough are 10-12ug/ml and 1-2ug/ml, respec­tively. The blue line shows the appro­pri­ate admin­is­tered dose while the black line shows one that is excessive.

 

 

The route of admin­is­tra­tion is deter­mined by the site of infec­tion, the level of antibi­otic required at that site, and the patients sta­tus and the urgency of appro­pri­ate treatment.

• Intra­venous (IV) admin­is­tra­tion typ­i­cally gives the high­est con­cen­tra­tions in the short­est time and is thus pre­ferred for seri­ously ill patients
• Intra­mus­cu­lar (IM) admin­is­tra­tion pro­duces lower con­cen­tra­tion lev­els over a longer period than IV but a higher con­cen­tra­tion than oral administration.
• Oral admin­is­tra­tion is ben­e­fi­cial, depen­dent on the infec­tion, for main­te­nance doses.
• Other meth­ods of admin­is­tra­tion include sub­lin­gual, muco­cu­ta­neous (such as eye, nose, ear) and topical.

For seri­ously ill patient with SIRS and a gram pos­i­tive gram stain from the C&S, what route of antibi­otic admin­is­tra­tion do you recommend?

IV because it’s the fastest possible.

Bioavail­abil­ity affects the con­cen­tra­tion of avail­abil­ity of the active drug in the sys­temic cir­cu­la­tion because the drug must suc­cumb to the gas­troin­testi­nal tract. It’s also influ­enced by sol­u­bil­ity, which relates to the excip­i­ents (vehi­cle; tablets and cap­sules for oral, sup­pos­i­to­ries for rec­tal) in which the mate­r­ial is in. In regards to the ques­tion of gener­ics, the FDA reg­u­lates the gener­ics so they are the same as the pro­pri­etary ver­sions (at least for those man­u­fac­tured in the USA).

An exam­ple of bioavail­abil­ity are the Flu­o­ro­quinolones (Ciprofloxacin, Lev­ofloxacin and so forth). It’s rec­om­mended that you should avoid tak­ing it 2 hours before a meal or 4 hours after a meal and also avoid antacids.

LOOK AT PAGE 493 FIGURE 21.3.

Drug dis­tri­b­u­tion in body tis­sues should be con­sid­ered in pharmacology/pharmacokinetics. IM/IV/Oral, it doesn’t mat­ter, the drugs all get to the blood/systemic cir­cu­la­tion. Even the nasal cav­i­ties are highly vas­cu­lar and it has been shown (chi­nese showed this for cen­turies, long before jen­ner for­mu­lated it). Some­times, you want the drug to be con­cen­trated in the lungs some­times, but with a pill taken by mouth, it has to cross into the blood stream. If it’s bound to pro­tein, it can’t cross because the sheer size of it pro­hibits cross­ing. This is some­thing the drug man­u­fac­turer has to doc­u­ment that pro­tein bind­ing is not sig­nif­i­cant or there is some mech­a­nism where the drug can be freed by the protein.

If the drug binds to pro­tein it will not cross into the tis­sue. (In other words, protein-bind = can­not cross)

Not pro­tein bound (unbound drug) means it crosses by…

1. dis­solv­ing in lipid (lipid sol­u­ble drugs) because the cell mem­branes are made of lipid.
2. pro­tein pores which are trans­port proteins
3. means of enzyme cat­alyzed transport

 

One of the quin­tes­sen­tial anti­fun­gals: Ampho­tericin B, used widely for sys­temic fun­gal infec­tions, is extremely toxic. And why would that be an issue? Both fungi and your cells are eukary­otic. In the busi­ness this is known as amphoter­ri­ble. Some patients rather die than take it because if they do take it they get this metal­lic taste in their mouth, per­sis­tent ear ring­ing, nau­sea, upset GI and vomiting.

Part of the tox­i­c­ity can be over­come and reduced by tar­get­ing deliv­ery. Ampho­tericin B is now encap­su­lated in lipo­somes (now known by the mar­ket name: ambi­some). A lipo­some is a fat glob­ule (lip=lipid=fat), and inside of it is ampho­tericin B. It will dis­solve in the lipid mem­brane due to the sim­i­lar struc­ture and that will ensure that the deliv­ery to the tis­sues is much more facile rather than cir­cu­lat­ing ad libi­tum (freely) through­out the body. Ampho­tericin B is now mar­keted as Ambi­some because it’s in this lipid vesicle.

Ref­er­ence Page 494: Read “Drug Dis­tri­b­u­tion” sec­tion to get a fuller idea.

Ciprofloxacin has an extended prepa­ra­tion of cipro. There are sev­eral coat­ings so there’s an extended release of the drug (by dis­solv­ing the lay­ers slowly), that allows a dif­fer­ent dos­ing regimen.

Spec­trum of activity

Antibi­otic: Only refers to drugs that are effec­tive against bacteria.

Antimi­cro­bial: Includes antipar­a­sitic, anti­fun­gal, antivi­ral. An antivi­ral drug is com­monly referred to as an antibi­otic but it’s not an antibiotic.

The range refers to the num­ber and vari­ety of organ­isms that can be cov­ered. When we talk about any antimi­cro­bial agent, we talk about whether it’s a broad spec­trum (cov­ers a wide range of organ­isms) or nar­row spec­trum (cov­ers a nar­row range of organ­isms; gen­er­ally one type of bacteria).

Turn to page 495, fig­ure 21.5: YOU MUST KNOW THIS CHART. Know which antibi­otic is used against what type of bacteria

Pen­cillin: When asked for a nar­row spec­trum antibi­otic on the test, DO NOT say peni­cillin, as it can be used against out­side of its spec­trum of gram pos­i­tive activ­ity, even though the table in the book says it’s a nar­row spec­trum. Ampi­cillin is a vari­a­tion of peni­cillin. Up to 15–20 years ago, peni­cillin was the treat­ment of choice for N. gon­or­rhoea and it’s not any­more due to resistance.

Isoniazid/Ethambutol: Nar­row spec­trum, effec­tive against mycobacteria.

Polymyxin B: Nar­row spec­trum: effec­tive against gram neg­a­tive bacteria.

Broad spec­trum:

• Ampi­cillin (cov­ers gram pos­i­tive and gram negative)
• Chloramphenicol/tetraycycline (cov­ers every­thing except mycobacteria)
• Strep­to­mycin (cov­ers every­thing except gram positive)

What is the sig­nif­i­cance of using broad ver­sus narrow?

1. Broad spec­trum gives greater per­tur­ba­tions to nor­mal flora so it leads to sec­ondary infec­tions (pseudomem­bra­nous col­i­tis; antibi­otic asso­ci­ated colitis).

2. The pos­si­bil­ity of resis­tance will be increased with broad spec­trum because it’s used against a wider vari­ety of organ­isms. In other words, the chances of resis­tance devel­op­ing are more likely with the broad spec­trum antibi­otic because it cov­ers such a wide range of organ­isms so there’s a greater poten­tial for resistance.

Some antibi­otics are clas­si­cally asso­ci­ated with this: Clin­domycin, for exam­ple, in the early days had an unsa­vory rep­u­ta­tion for an increased inci­dence for pseudomem­bra­nous col­i­tis and it was par­tic­u­larly use­ful against anaer­o­bic bac­te­ria. The broader the spec­trum, the more likely it is to per­turb nor­mal flora.

You will be asked a ques­tion on fecal trans­plant (aka fecal bacteriotherapy):

• Fecal trans­plant restores the colonic flora by intro­duc­ing healthy bacteria.
• The pro­ce­dure involves sin­gle to mul­ti­ple infu­sions (by enema, colono­scope, etc) of bac­te­r­ial fecal flora from a healthy donor, into a patient suf­fer­ing from Clostrid­ium dif­fi­cile infec­tion (pseudomem­bra­nous col­i­tis; antibi­otic asso­ci­ated colitis).

Resis­tance to Antibiotics/Antimicrobial agents

Fac­tors influ­enc­ing resistance:

1. Mis­use
2. Med­ical “Mal­prac­tice” (overuse). Not delib­er­ate but an overuse.
3. Cross bor­der / Inter­net access / Inter­na­tional access with­out prescription.
4. Ques­tion­able formulations(preparation off­shore) / Counterfeits
5. Self med­ica­tion
6. Ram­pant & aggres­sive adver­tis­ing by “BIG pharma.”
7. Increased patient demand and access.
8. FDA pres­sures. (Not so com­mon with antibi­otics but rich young white pro­fes­sion­als, like doc­tors, lawyers, pres­sure the FDA to fast track the med­ica­tions w/o ade­quate research and the data may be soft.)
9. Antibi­otic residues in meat/fish. Xeno­bi­otics can be hormonal-like, and in some coun­tries like Brazil you have early onset of puberty.
10. Ani­mal hus­bandry. (sup­ple­ment­ing cat­tle, feed with antibi­otics & hormones..etc. Cat­tle put on more weight and they can make more money)
11. Poor patient com­pli­ance (Pre­ma­ture dis­con­tin­u­a­tion or miss­ing a dose; Cer­tain lev­els are required for efficacy)
12. Access and availability
13. Insur­ance cov­er­age, which also affects avail­abil­ity. (It’s not your doc­tor that decides what drug you get, it’s your insur­ance). A social disaster.
14. Migra­tion / Inter­na­tional Travel / Off­shore med­ical treat­ments. The lat­est denizen on the block, Extended spec­trum organisms.
15. Med­ical tourism (TNA: tit­ties and ass in India).
16. Con­tin­ued adap­tive responses, muta­tions, changes in the tar­get organism.

Bac­te­ria can adapt so quickly to changes, it sur­passes any­thing we are capa­ble of. Every sin­gle antibi­otic dis­cov­ered to date has been shown to be asso­ci­ated with resis­tance in tar­get organ­isms, even in cir­cum­stances where there is no evi­dence of prior exposure.

Since bac­te­ria have such a won­der­ful adap­tive capa­bil­ity and can mul­ti­ply so rapidly, that one in a mil­lion or bil­lion chance cre­ates nat­u­rally occur­ring resis­tant organ­isms. You start off with a mixed pop­u­la­tion of resis­tant and sus­cep­ti­ble organ­isms. You treat with antibi­otics and the sus­cep­ti­ble ones will be elim­i­nated and you’re left with a resis­tant pop­u­la­tion which will mul­ti­ply like crazy and that drug no longer become effective.

For this rea­son, the CDC launched an antibi­otic track­ing sys­tem for hos­pi­tals. Antibi­otic use can lead to antibi­otic resis­tance which can increase the risk of hos­pi­tal­iza­tion or lead to treat­ment fail­ure in sick patients, accord­ing to the CDC.

Antibiotic-resistant infec­tions are likely to require longer and more costly hos­pi­tal stays and have higher mor­tal­ity rates. If first-line drugs don’t clear up the infec­tion, sec­ond or third-line drugs may be even less effec­tive, more toxic, and more expensive.

Real threat of return­ing to a pre-antibiotic era: Drugs no longer use­ful because organ­isms are resis­tant to every­thing we have come up with.

With every sin­gle antibi­otic noted to date, within 6 months of use, antibi­otic resis­tance has been doc­u­mented. So there’s no antibi­otic which bac­te­ria have not devel­oped resis­tance. It’s micro­bial sur­vival of the fittest.

*T/F Ques­tion! - Emerg­ing resis­tance to N. Gonnorhea is devel­op­ing. (true)

Basic mech­a­nisms of resistance

One sin­gle organ­ism may have sev­eral dif­fer­ent mech­a­nisms to sev­eral drugs. The bac­te­ria can either destroy the drug or alter the drug or inhibit the drug or do all of them.

Ref­er­ence: pg. 496 Table 21.2

This isn’t an exhaus­tive list, there’s many vari­a­tions of each theme. Know the 3 pri­mary mech­a­nisms and one antibi­otic asso­ci­ated with each one. These are all ways that bac­te­ria can destroy/alter/inhibit the drug (antibiotic).

1. Destroys. There are sev­eral ways.
   1. Plas­mids can encode enzymes that chem­i­cally alter amino­gly­co­sides (by acety­la­tion or phosphorlyation)
   2. Plas­mids can encode beta-lactamase which inac­ti­vate beta-lactam antibi­otics such as peni­cillin and cephalosporins.
2. Alters drug.
   1. An amino­gly­co­side nor­mally tries to bind to a 30S ribo­some sub­unit, so as a resis­tance mech­a­nism, the bac­te­ria makes a dif­fer­ent, altered ribo­some that won’t allow the drug to bind to it and won’t be inhibited.
   2. Quinolone nor­mally tries to bind to DNA topoi­so­merase to inhibit DNA syn­the­sis but the bac­te­ria could make an altered DNA topoi­so­merase so the antibi­otic won’t bind to it.
   3. Rifampin nor­mally binds to RNA poly­merase to inhibit RNA syn­the­sis but the bac­te­ria makes an altered poly­merase so it won’t bind to the drug.
3. Inhibits drug entry or removes drug.
   1. Peni­cillin binds to pro­teins in the out­er­mem­brane to enter the cell and block pep­ti­do­gly­can syn­the­sis, but the bac­te­ria can change the shape of these porin pro­teins so the drug can’t enter.
   2. Ery­thromycin and tetra­cy­cline nor­mally bind to ribo­some sub­units to inhibit pro­tein syn­the­sis, but the bac­te­ria cre­ates a new mem­brane trans­port sys­tem to lit­er­ally pump the drug out of the cell.

Increas­ing the num­ber of dif­fer­ent mech­a­nisms of resis­tance increases the like­li­hood of resistance.

T/F Ques­tion – Bac­te­ria are con­scious or sen­tient enti­ties. (true)

What evi­dence is there that bac­te­ria are con­scious or sen­tient entities?

Cap­sule, spore , nutri­tion, tem­per­a­ture, adap­tive, lis­te­ria cold enrichment.

1. Cap­sule for­ma­tion is deter­mined on nutri­ent avail­abil­ity. Bac­te­ria respond to nutri­ent excess by mak­ing a cap­sule which is pro­tec­tive and allows them to attach to tis­sues to pro­duce infection.
2. Spore for­ma­tion and ger­mi­na­tion are dependent/determined by envi­ron­ment. When nutri­ents are lim­ited and water is not avail­able, they will form spores. When con­di­tions are favor­able, they will germinate.
3. Motil­ity asso­ci­ated with fla­gella. Taxis (move­ment) is a direc­tional response. Pho­to­taxis. Mag­ne­to­taxis. These are all responses to var­i­ous stim­uli which means they need to be coör­di­nated to move in a cer­tain direction.
4. Quo­rum sens­ing (we talked about this with biofilms!)

Eval­u­a­tion of Efficacy

How do you know if antibi­otics are going to work? It’s the S in C&S (Sensitivity).

Sus­cep­ti­bil­ity test­ing (for lab practicum).

1. Kirby Bauer Disc diffusion
   1. R – not effective
   2. I – when there’s evi­dence of accu­mu­la­tion of organ­ism in blood/bodily fluids.
   3. S – use
2. MIC, MBC -> Broth Dilution
3. Agar Dilu­tion
4. “E” Test

Ques­tion on exam You have a plate with antibi­otics, and you look for the zone of inhi­bi­tion. This zone is arbi­trary and has no sig­nif­i­cance until you refer to the NCLS Chart. If the chart says that 12mm or more is equiv­a­lent to sus­cep­ti­bil­ity, whereas 10 or less is resis­tant, and you have 12 or more, it means it’s sus­cep­ti­ble and so forth

If con­trol zones are not within accept­able lim­its or if the iden­tity of the organ­ism is unknown, the data from the test is invalid and must be dis­carded neces­si­tat­ing that the test be repeated.

A high M.I.C. (min­i­mum inhi­bi­tion con­cen­tra­tion) typ­i­cally indi­cates resis­tance. (on the KB chart) I’ll give you zone sizes, the results of the test and ask you whether it’s R or S or I

Now turn to pg. 500 – dilu­tion of antibiotics.

You have dif­fer­ent graded con­cen­tra­tions, all tubes inoc­u­lated with the same organ­ism. The tubes with no antibi­otics are con­trol. If the con­trol tubes have no growth, they are dead. (HE WILL ASK THIS). 2 mg/mL has no growth, so the MIC is the low­est con­cen­tra­tion that inhibits the growth of the bac­te­ria. I could give you the results like this or a chart and ask you where the MIC is.

To deter­mine the bac­te­ri­ci­dal con­cen­tra­tion, take all the neg­a­tive tubes and put them on plates. If they grow, they are not totally inhib­ited. The MBC in this case is 8 mg/ml. The MBC is the min­i­mum bac­te­ri­ci­dal con­cen­tra­tion: the low­est con­cen­tra­tion that KILLS the organ­ism, estab­lished by tak­ing cul­tures with no obvi­ous growth, to antibiotic-free media and look­ing for growth. If the sub­cul­ture in the antibiotic-free media grows, the bac­te­ria were inhib­ited but not killed; The MBC is equated by those neg­a­tive cul­tures that do not grow on the antibiotic-free media, doc­u­ment­ing that the bac­te­ria were killed.
Uses/Applications of MIC/MBC Data: This is a guide to the choice of therapy.

1. If the MBC 64x times greater than the MIC, that indi­cates TOLERANCE which means the antibi­otic is not likely to be effec­tive for treat­ment. You have to use so much of the antibi­otic to kill the organ­ism that you exceed the ther­a­peu­tic ratio or range and you can harm the patient.

2. Effi­cacy is manda­tory data for FDA approval and essen­tial documentation.

3. Com­par­a­tive effi­cacy. In other words, the FDA is not nec­es­sar­ily con­cerned with prov­ing just another drug with­out a sig­nif­i­cant advan­tage, unless it’s a generic, which the data would have been estab­lished by the pro­pri­etary brand name. So this is one of the major planks in the approval process so they have to demon­strate that the drug actu­ally works.

4. Even more impor­tantly, it mon­i­tors the devel­op­ment of resis­tance, or changes in sus­cep­ti­bil­ity patterns/response.

 

You’ll typ­i­cally see:

MIC₅₀ means ½ of the iso­lates tested are inhib­ited (which means they are susceptible).

MIC₉₀ means 90% (9÷10) are inhibited.

MBC₅₀ means ½ of the iso­lates are killed by the concentration.

MBC₉₀ means 90% of the iso­lates are killed by the concentration.

YOU’LL NEVER SEE MBC 100. There’s always going to be an organ­ism that is going to be resis­tant, given the fact that antibi­otic resis­tant can be doc­u­mented with­out prior expo­sure and within 6 months of intro­duc­tion of any antibiotic.

If you require an increase of con­cen­tra­tion, it’s indica­tive of an increase in resis­tance, you need a greater con­cen­tra­tion of antibiotics.

↑ [C] = ↑[R] (C=concentration, R=resistance)

In the year 2000, MIC₅₀ = 1ug/ml

In the year 2010, MIC₅₀ = 5ug/ml

Let’s look at ciprofloxacin… If an effec­tive dose is achieved at 5ug/ml, and you have MIC₅₀ but years down the line 50% of your iso­lates may require 10ug/ml.  If you have an organ­ism that requires TWICE the con­cen­tra­tion to be inhib­ited, it means 5ug/ml is not going to be effec­tive any­more. You can’t just dou­ble the dose cause then you have toxicity.

Be famil­iar with why an organ­ism show­ing tol­er­ance is not effec­tive to that treatment.

Agar dilu­tion: Rather than hav­ing a bunch of tubes for the MIC/MBC, the tubes are replaced by agar, with vary­ing con­cen­tra­tions of antibi­otic or antimi­cro­bial agent.

Neu­tropen­ics. The big prob­lem is neu­tropen­ics are treated with anti­fun­gal treat­ments like Nys­tatin Px but a lot of those patients end up with fatal fun­gal infec­tions. So the ques­tion is… are the deaths due to anti­fun­gal fail­ure? In other words, were the organ­isms resis­tant to nys­tatin? The fail­ure of treat­ment was not due to the resis­tance of the organ­isms.. none of the fungi were resis­tance. The fail­ure was due to the neu­trophil count not being increased.

Fungi are not tested rou­tinely for sus­cep­ti­bil­i­ties, although sev­eral experts are rec­om­mend­ing that this be done. For the most part, the responses are pre­dictable. What is done instead is the tests are done in pools of iso­lates and they are peri­od­i­cally tested with the agar dilu­tion or now they have mod­i­fi­ca­tions with respect to instru­men­ta­tions that allow you to detect growth depen­dent on the concentrations

E test

 

The E test is not as widely accepted in Amer­ica because here’s a bit of jin­go­ism since it wasn’t devel­oped in the USA. It’s slowly gain­ing accep­tance and becom­ing more rou­tine now.

The E test is use­ful because you have a lawn of bac­te­ria and you could put sev­eral dif­fer­ent antibi­otic strips that could be van­comycin, peni­cillin, gen­tamycin, tobramycin. Who­ever came up with this is cer­ti­fi­ably genius.

The con­cen­tra­tions of the antibi­otics are lay­ered sequen­tially and sold com­mer­cially. You put this strip on the lawn and you end with growth or no growth or a zone of inhibition.

The point of inter­sec­tion where the point of inhi­bi­tion coin­cides with the level marked on the strip is the MIC. The beauty of this is you could test 4–5 dif­fer­ent antibi­otics sep­a­rately and you could get break-point MICs. You will indu­bitably get ques­tions on the exams with this.

 

The E test, a gra­di­ent dif­fu­sion method that deter­mines antibi­otic sen­si­tiv­ity and esti­mates min­i­mal inhibitory con­cen­tra­tion. The plas­tic strip, which is placed on an agar sur­face inoc­u­lated with test bac­te­ria, con­tains an increas­ing gra­di­ent of the antibi­otic. The MIC in ug/ml is clearly shown.

Sites of Action and Exam­ples of Antibi­otics Tar­get­ing PROKARYOTES

Ref­er­ence page 501, fig­ure 21.10

You see prokary­otes and eukary­otes. There’s a clin­i­cally sig­nif­i­cant dif­fer­ence between the two. This is a prime exam­ple. Not only do these dif­fer­ences reflect speci­ficity of action but explain the tox­i­c­ity asso­ci­ated with antimi­cro­bial agents.

How is tox­i­c­ity of antibi­otics and adverse drug reac­tions related to the dif­fer­ences of eukary­otes and prokaryotes?

The speci­ficity of micro­bial action is related to the struc­tural, phys­i­o­log­i­cal and meta­bolic dif­fer­ences that dif­fer­en­ti­ate microbes from the human body.

You must be famil­iar with ALL of these sites and the rel­e­vant antibi­otics that tar­get those spe­cific sites. Any part of the bac­te­ria is fair-game.

What antibi­otics inhibit the bac­te­r­ial cell wall?

• Peni­cillin
• Cephalosporins
• Van­comycin
• Bac­i­tracin (used top­i­cally as an antibiotic).

Read the “Cell Wall” para­graph. You only find pep­ti­do­gly­can in bac­te­ria. These agents have no effect on eukary­otic cells because all eukary­otic cells lack peptidoglycan.

Ques­tion on test: What is the treat­ment of choice for L forms and why? An agent that inhibits pro­tein syn­the­sis (gen­tamycin) and peni­cillin that inhibits the ref­or­ma­tion of the cell wall. Some L forms can reform the wall, so if they reform the wall, you use peni­cillin to inhibit the cell wall and the gen­tamycin to inhibit pro­tein syn­the­sis, so that’s a syn­er­gis­tic exam­ple of antibi­otics. (Syn­ergy = 1+1=11).

Beta-lactam ring -> Peni­cil­li­nase (beta-lactamase) -> Pro­duces peni­cil­loic acid, which inac­ti­vates the peni­cillin. The peni­cil­loic acid is also a prod­uct that can trig­ger allergic/hypersensitive reactions.

What antibi­otics inhibit the bac­te­r­ial cell membrane?

• Polymyxin B. You don’t use it as a wide­spread use of an antibi­otic. It’s used top­i­cally. Read “Cell Mem­branes” para­graph. ALL cells have mem­branes so it’s not an ideal tar­get of drug action but there are some dif­fer­ences between micro­bial and mam­malian cell mem­branes. Most drugs that tar­get cell mem­branes are highly toxic when admin­is­tered systemically.
• What is the basis of speci­ficity of polymyxin? The eukary­otic cell mem­brane has a dif­fer­ent sterol com­po­si­tion that isn’t rec­og­nized as well by the polymyxin.

What antibi­otics inhibit the Nucleic Acid synthesis?

• Rifampin (RNA)
• Quinolones (DNA)
  • FQ = Flu­o­ro­quinolones. ie.,
    • Ciprofloxacin
    • Lev­ofloxacin
    • Temafloxacin
    • When you see FLOXACIN in the drug word, that’s a Fluoro­quinolone and they inhibit DNA synthesis
  • What is the basis of action of FQ’s? How do they act? They inhibit an enzyme that is cen­tral to DNA repli­ca­tion that ini­ti­ates and causes neg­a­tive super coil­ing to the DNA prior to split­ting by heli­case, called DNA gyrase aka topoi­so­merase. Both human and bac­te­ria require DNA gyrase but there’s Type II topoi­so­merase that’s found in prokary­otes. The bac­te­r­ial DNA gyrase (enzyme) dif­fers from the eukary­otic DNA gyrase.
  • The stu­pid way I remem­ber flu­o­ro­quinolones inhibits TOPOISOMERASE activ­ity… is that they are both big words with many syllables! 
  • The FQ is going to pref­er­en­tially bind to the bac­te­r­ial enzyme to a much lesser extent than the human gyrase.
  • The major adverse effect of FQ is arthropa­thy (joint pain), pri­mar­ily asso­ci­ated with a prin­ci­pal weight bear­ing joint because it inhibits the con­ver­sion of car­ti­lage to bone. That is why chil­dren are most sus­cep­ti­ble to this adverse effect but adults will get ten­donitis, ten­don rup­ture, bur­si­tis as well.
  • Another adverse effect is that it causes pho­to­sen­si­tiv­ity and pho­to­mu­ta­ge­n­e­sis which are light-induced changes to DNA.

 

The prod­uct mono­graph is a qua­si­le­gal guide­line doc­u­ment. As a patient you might get a pack­age insert that gives you the adverse side effects and how you should take the dose and what­ever else. First thing is shows is the warn­ing. FQ’s are asso­ci­ated with an increased risk of ten­dini­tis and ten­don rup­ture (Is your ACL a ten­don or lig­a­ment? It’s a lig­a­ment. Ten­dons are mus­cle to bone. What are lig­a­ments? Bone to bone con­nec­tive tis­sue. What’s the sim­i­lar­ity? They’re both dense con­nec­tive tissue).

• Pharmacology/Pharmakinetics tells you how the drug is dis­trib­uted through­out the body which tells you the effi­cacy. The bind­ing of cipro to serum pro­teins is 20–40% which isn’t likely to be high enough to cause sig­nif­i­cant pro­tein bind­ing inter­ac­tions w/ other drugs.

 

What antibi­otics inhibit Pro­tein synthesis?

• Amino­gly­co­sides (Pro­teins are made of amino acids, so aminogly­co­sides inhibit pro­tein synthesis)
  • Tobramycin
  • Gen­tamycin
  • Where have we seen gen­tamycin men­tioned before?
  • Gen­tamycin + Peni­cillin = syn­er­gis­tic effect against L forms.
  • Gen­tamycin peak and trough = 10-12ug/ml and 1-2ug/ml
  • peak is reached in 2 hours, trough in 4 hours. (symmetrical)
  • side effects if too much: oto­tox­i­c­ity and nephrotoxicity

These antibi­otics require mon­i­tor­ing. The ther­a­peu­tic accept­able peak lev­els of these are 10-12ug/ml and the trough is 1-2ug/ml. The peak is usu­ally 2 hours and the trough is 4 hours after admin­is­tra­tion. Peak and trough lev­els are a guide to the effec­tive lev­els which are not to be exceeded to avoid tox­i­c­ity. This is par­tic­u­larly impor­tant in patients with defi­cient kid­ney func­tion. What would you do if lev­els exceeded peak and trough? You could lengthen the dosage fre­quency or lower the dose but you can’t arbi­trar­ily do that because you have to con­sider the con­cen­tra­tion achieved at the site of infec­tion. If you need 5ug/ml in the CSF and you’re only get­ting 2ug/ml, is it going to be effec­tive? NO. So you need to con­sider MIC, MBC and the con­cen­tra­tion achieved at the site of infection.

Side effects: Oto­tox­i­c­ity and nephro­tox­i­c­ity (mid­dle ear dam­age and kid­ney dam­age) are the two pri­mary con­se­quences of amino­gly­co­side if peak and trough lev­els are exceeded, espe­cially in elderly.

• Chlo­ram­pheni­col (page 509, fig­ure 21.14)
• Chlo­ram­pheni­col is a BROAD SPECTRUM antibi­otic! (review page 495 figure)
• In rare cases, chlo­ram­pheni­col is fatal because it can cause aplas­tic ane­mia (the bone mar­row com­pletely stops pro­duc­ing blood cells). This hap­pens in 1 out of 30,000 drug recip­i­ents and is inevitably fatal.
• Another threat is gray baby syn­drome because new­borns may die from mul­ti­ple toxic com­pli­ca­tions so this drug is ONLY given to extremely ill hos­pi­tal­ized patients.
• Tetra­cy­clines
• Ery­thromycin

 

What antibi­otics inhibit Folic Acid Syn­the­sis? (page 502)

1. sul­fon­amides
2. trimethoprim/sulfamethoxazole
   1. Exam­ples: Bactrim or Septra,
   2. http://en.wikipedia.org/wiki/Trimetho­prim/sul­famethox­a­zole

In this case we are look­ing at metab­o­lism. FA is a pre­cur­sor to the biosyn­the­sis of nucleic acids. Trimetho­prim blocks the path­way in which folic acid is con­verted into nucleic acids in bac­te­ria but not in humans because humans use a dif­fer­ent path­way to uti­lize that folic acid.

UTI: One of the most widely used antibi­otics for treat­ment of UTI. Where do the bac­te­ria pri­mar­ily come from that cause UTI? The area between the gen­i­talia and the rec­tum. Also, hon­ey­moon cys­ti­tis is exces­sive sex­ual inter­course which pre­dis­poses to UTI.

Cran­berry juice does not treat UTI’s, it reduces the inci­dence rate.

Pneu­mo­cys­to­sis / Pneu­mo­cys­tic pneu­mo­nia is a form of pneu­mo­nia caused by a yeast-like fungus.

 

Sites of Action and Exam­ples of Antibi­otics Tar­get­ing EUKARYOTES (Fungi, Pro­to­zoa, Worms)

Folic acid synthesis

• trimetho­prim (same as prokaryotes)

Nucleic acid synthesis

• flucy­to­sine (RNA) an anti-fungal agent (coin­ci­den­tally fluoro­quinolones inhibit nucleic acid syn­the­sis [DNA though] in prokary­otes, they both start with fl… hope­fully that helps)

Cell mem­brane:

• ampho­tericin B
• imi­da­zoles
• nys­tatin

Anti­fun­gal drugs (pg 511)

• nys­tatin
• ampho­tericin b
• imi­da­zole and triazole
• flucy­to­sine
• grise­o­ful­vin

Antipar­a­sitic drugs (pg 512)

• meben­da­zole
• metron­ida­zole

In Table 21.5: Remem­ber Malarone for Malaria:

Malarone is the syn­er­gis­tic com­bi­na­tion of 2 drugs: ato­vaquone and proguanil (he will prob­a­bly ask this!)

Antivi­ral drugs (page 514)

• Acy­clovir inhibits viral DNA
• Rib­avirin inhibits viral RNA

Know how Acy­clovir works!

Acy­clovir is struc­turally sim­i­lar to the nucleotide guano­sine. An enzyme, found only in virus-infected cells con­verts acy­clovir to acy­clovir triphos­phate, the actual antivi­ral drug. This false nucleotide com­petes with GTP to incor­po­rate into DNA, lead­ing to chain ter­mi­na­tion and stops DNA replication.

REVIEW

What is the major con­traindi­ca­tion asso­ci­ated with antibi­otic use? An indi­ca­tion is a cir­cum­stance of an infec­tious dis­ease with char­ac­ter­is­tic eti­ol­ogy that war­rants use of an antibi­otic. Con­traindi­ca­tions pre­clude to negate indications.

1. A his­tory of aller­gic response or hyper­sen­si­tiv­ity is a con­traindi­ca­tion, includ­ing ana­phy­lac­tic shock which can be fatal.

2. Syn­ergy: an ampli­fied effect beyond an addi­tive. The sum of the parts are more effec­tive than their indi­vid­ual selves (1 plus 1 = 11). (malarone = ato­vaquone and proguanil syn­er­gis­tic combination)

Toxic effects of fluoroquinolones?

Arthro­topy, rup­ture of the ACL, ten­dini­tis, bur­si­tis, pho­to­sen­si­tiv­ity, photo-mutagenicity. The major prob­lem is arthro­topy in every age group but par­tic­u­larly chil­dren. While you’re on flu­o­ro­quinolones, you have to avoid direct light.

———————

ESBL = Extended spec­trum beta-lactamase bac­te­ria (beta lac­ta­mase 2.0)

With ESBL’s, if the organ­ism is resis­tant to peni­cillin then it’s going to be resis­tance to beta lac­tam antibi­otics because it means it has beta lactamase.

“The dev­il­ishly clever bac­te­ria met our chal­lenge by cre­at­ing beta lac­ta­mase, an enzyme that grants many bac­te­ria immu­nity to penicillin-type antibi­otics. In turn, we upped the ante by devel­op­ing new kinds of antibi­otics that trounced these beta lactamase-producing pathogens.

But the bac­te­ria weren’t done yet. Some tricky lit­tle bugs had a trick up their metaphor­i­cal sleeves: Beta Lac­ta­mase model 2.0, known to us as extended-spectrum beta lac­ta­mase, or ESBL. This enzyme not only chops apart peni­cillins, but cephalosporin antibi­otics, too (all of the antibi­otics whose generic names begin with “Cef-”).”

Page 278… Table 1.4, Know the prob­lems asso­ci­ated with atyp­i­cal bacteria.

The part about chlamy­dia.. they are atyp­i­cal because they don’t have a cell wall … you can’t grow chlamy­dia on blood agar, it is an oblig­ate intra­cel­lu­lar pathogen so it must grow on tis­sue culture.

What are the 3 species of chlamy­dia that infect humans? Chlamy­dia pneu­mo­niae, tra­choma­tis (blind­ness, STD), and psittaci (causes ornitho­sis or psit­ta­co­sis; par­rot fever)

Chlamy­dia pneu­mo­nia. What is the pri­mary sig­nif­i­cance? It causes pneu­mo­nia! Sec­ondary sig­nif­i­cance: Coro­nary artery dis­ease; myocar­dial infarc­tion due to high anti­body lev­els. Peo­ple who have had heart attacks have high lev­els of anti­bod­ies which means they have been exposed to the bacteria.

Rick­ettsia: What is the only rick­ettsia that is able to sur­vive out­side the liv­ing host? Cox­iella bur­netii is the only rick­ettsia that could sur­vive out­side of the liv­ing host. It forms a spore like entity which isn’t equal to the bac­te­r­ial endospore but can sur­vive in soil, feces, etc for an extended period of time. It’s the only rick­ettsia that can do that and the eti­ol­ogy of Q fever.

Microscopy: Know the lim­its of res­o­lu­tion, the var­i­ous types of micro­scopes (bright­field is the most com­mon, elec­tron microscopy is used for viruses, flu­o­res­cent is used for iden­ti­fy­ing antigens/antibodies and you don’t need iso­la­tion in pure cul­ture to do flu­o­res­cent marking).

• Human eye: 200 micrometers.
• Bright­field: 200 nanome­ters (0.2 micrometer)
• Scan­ning: 10–20 nanometers
• Trans­mis­sion = 1–2 nanometers.

You will be asked some­thing like: How many times smaller of an object can be seen with a scan­ning elec­tron micro­scope than the naked eye? Con­vert every­thing to nanome­ters to find out.

Review pre­vi­ous tests. Review those god damned pre­sen­ta­tion sheets. Review fuck­ing every­thing. Good luck.