Antimicrobial   therapy
MUDr. Lenka Černohorská, Ph.D.
nAntibiotics are substances against  bacteria
n
nOther groups:
nAntivirotics – against viruses
nAntituberculotics - against mycobacteria
nAntiparasitics – against parasites
n
Targocid

Antibiotics are devided due to mechanism of efficacy into 4 groups:
1.Inhibition of cell wall synthesis (betalactames, glycopeptides)
2.Cell membrane destroy (polypeptides)
3.Inhibition of NA syntesis (quinolons, imidazols)
4.Inhibition of proteosyntesis (tetracyclines, chloramphenicol, macrolides, lincosamides,
aminoglycosides)
* Attack against bacterial metabolism (sulfonamids)
1.

Betalactames
* Baktericidal, only for growing bacteria
* Often causes allergy
•
nPenicillins (PNC, oxacillin, ampicillin, piperacillin)
nCefalosporines (1.- 4. generation)
nMonobactams (aztreonam)
nCarbapenems (imipenem, meropenem)

Glycopeptides
* Reserved for G+ bacteria
* Vancomycin and less toxic, but more expensive teicoplanin
•
•Polypeptides
* Ototoxic and nefrotoxic
* Polymyxin B only local as part of ear drops -  Otosporin
* Polymyxin E – colistin rare used
* Primary resistence: all G+ bacteria, proteus, providencia, morganella, serratia etc.
*
N

Aminoglycosides
* Bactericidal, ototoxicity and nefrotoxicity
* Synergy with betalactames – decrease of toxicity
* Preparates: Streptomycin only against tuberculosis, gentamicin, netilmicin, amikacin, neomycin
with bacitracin = framykoin (neomycin is too toxic, only for local using)

•Tetracyklines
•
* Broad spectrum
* Don‘t use until 10 years (teeth development)
* Less used
*
•Chloramphenikol
* Broad spectrum
* Good penetration to liquor, Hematotoxicity
*
•
t

Macrolides
•I. generation: erythromycin, rare used
•II. generation: roxithromycin
•III. generation: clarithromycin, azithromycin – good intracellular penetration and longlasting
effect, for  G+ bacteria
•Lincosamides
* Lincomycin and clindamycin
* Reserved for surgery, good effect to G+ bacteria and anaerobes in addition to Clostridium
difficile – risc of pseudomembranous enterocolitis
•
•

Quinolones
* Bactericidal
* Don‘t use until 15 years (growth cartilages)
* I. generation (oxolin acid), II. generation (norfloxacin) only for urinary infection
* III. generation: ofloxacin, ciprofloxacin – also for systemic infection – often used
„
Ciphin250

Analogs of folate acid
* Sulfametoxazol in combination with trimetoprim form ko-trimoxazol known as BISEPTOL
* Bacteriostatic, worse penetrate into tissues
Nitrofurantoin (and nifuratel)
lEffectivity on sugar metabolism.  Bacteriostatic, broad spectrum
lFor urinary tract infection. Weighty undesirable effect: GIT disorder etc.
Other antibiotics
Linezolid (zyvoxid) – against resistant staphylococci

Nitroimidazols
* For anaerobes, for protozoas (T. vaginalis etc.)
* Metronidazol, Ornidazol
•
•Antituberculotics
* HRZS,HRZE - starting therapy (INH, rifampicin, pyrazinamid, streptomycin, etambutol) + other
* HRZ,HRE – sequenced therapy

Antivirotics
* Against herpes – acyclovir…
* CMV – gancyklovir, foscarnet
* Influenza – amantadin, rimantadin, tamiflu
* Antiretrovirus therapy – inhibitors of reverse transcriptase (nucleosid+nonucleosid) , inhibitors
of protease – in  combination
•Preparates: zidovudin, didanosin …
*

Antimycotics
* Fluconazol, itraconazol, ketoconazol  etc. – local (vaginal, skin infection)
* Amphotericin B – i.v. (in sepsis)
•
•Antiparasitics
* Against protozoa, helmintes, ectoparasites (moore in parasite capitol)
•
•Other preparates
* Antimalaric: primachin, chlorochin, meflochin…
* Leprosity: dapson
*

Susceptibility testing in vitro
*
*
* Do not correspondent in all cases with effect of therapy
* Quantitative tests (MIC, E-tests) – in  relevant patients
* Qualitative tests (disc diffusion method) – enough for common cases (susceptible - resistant)
*
*
*

Disc diffusion test
* MH agar is inoculated with suspension of bacteria
* Antibiotic discs (paper impregnated with antibiotics)
•are applied at MH – atb diffuse from disc through agar
* Concentracion of atb decrease with distance from disc
* If microb grow to disc/if there is little zone - is resistant (not susceptible)
* Big zone (higher than defined size) means susceptibility.
atbpsae21

Disc diffusion test
MH medium inoculated with bacteria
+ via dispensor aplied 6 papers impregnated with antibiotics
        Cultivation +
     (37°C/18-24h)
ATB diffuses from paper
R
s
R
Interpretation:
High zone= susceptible bacteria (S)
Small/Any zone=resistant bacteria (R)

Microdilution test (MIC)
* MIC is the lowest concentracion, which inhibites growth (first clear row)
* On paper stencil is asigned breakpoint. If MIC is lower than breakpoint, bacteria is susceptible.
If MIC is higher, bacteria will be resistent
* 1 plastic plate is used for 1 bacteria, for 12 antibiotics, in 8 various (decreasing)
concentracion (12th only in 7, because corner row upper right is growth control)

MIC – Material and methods
bacterial suspension
(0,5-1 CFU)
inoculation of plate with pipette
microtiter plate with 96 rows
cultivation (37°C/18-24h)
   reading
FAC468
1 row is a growth control

Interpretation
MIC I
MIC II
Case -susceptible
Case - resistant
4

PEN
AMS
CXT
CLI
CMP
MTR
PEN
AMS
CXT
CLI
CMP
MTR
4
64
128
32
64
64
4
64
128
32
64
KR
2
32
64
16
32
32
2
32
64
16
32
32
1
16
32
8
16
16
1
16
32
8
16
16
0.5
8
16
4
8
8
0.5
8
16
4
8
8
0.25
4
8
2
4
4
0.25
4
8
2
4
4
0.125
2
4
1
2
2
0.125
2
4
1
2
2
0.063
1
2
0.5
1
1
0.063
1
2
0.5
1
1
0.031
0.5
1
0.25
0.5
0.5
0.031
0.5
1
0.25
0.5
0.5
Interpretation of MIC - antibiogram – goes to clinician!
PEN (penicillin)….4……resistant
AMS (unasyn)……2…..susceptible

E-tests (quantitative)
* Similar to disc diffusion test, but strip is used
* An increasing concentracion of atb is used. Zone is egg like.
* There is a scale on strip – simply reading
etest
      MIC value is  0,75 mg/l (where borderline of zone cross the scale)

Resistance of microbes to antibiotics
* Primary resistance: all strains of bacteria are resistent.
* Secondary resistance: arises unsensitive mutants, by selective antibiotics pressure became
dominant
•*
•MBC (minimum bactericidal concentracion) is the lowest concentration, which kills bacteria
•Primary bactericid: atb, where MIC and MBC are almost equal
•Primary bacteriostatic: atb, where MBC is X-fold higher than MIC - unreal baktericidal effect in
human body
*

Resistance factors detection
* Special detection methods for resistance factors (for ex. betalactamase). It can be diagnostic
strips (chemical detection of specific ensym) or other tests (ESBL)
•1. Betalactamase testing
* In neisseria, M. catarrhalis, H. influenzae
* destroys betalactams
* For therapy we use ATB with inhibitors of betalactamase like clavulanate, sulbactam…
*
*

Detection of betalactamase
Paper with substrate
 + moisturing solution
Petri dish with bacteria
  Touch
Colour change (red)
After 30 sec red is missing
Reaction end (yellow)
Strain produces betalactamase

   2. ESBL (extended spectrum betalactamase)
•E. coli, K. pneumoniae etc. produces ESBL, which destroys cheap betalactams. For therapy we use
expensive carbapenems, aminoglycosides (toxicity), problem of ICU, big hospitals
•
•
•ESBL – screening
* Inhibition of growth between discs – owing to a synergism of 2-3 antibiotics such as aztreonam,
AMC, ceftriaxon
•
esbl
Amoxicilin/clavulanate
aztreonam

ESBL detection
* 4 discs: Cefotaxim (1) and ceftazidim (2), cefotaxim with clavulanate (3) and ceftazidim with
clavulanate (4)
* Difference between cefalosporines (1,2) and cefalosporines with clavulanate (3,4) is higher than
5mm
*
Mil02Pop
1
2
3
4
Compare
1 with 3 and 2 with 4