Control of Microbial Growth

Tim HoUniversity of Alberta, Canada* The materials are mostly based on Dr. Brian Lanoil’s Microb 265

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Part II

Objectives

1. Know 3 methods of microbial control

2. Know the strategies on how drugs control the growth of microorganisms.

3. Understand how do bacteria become resistant to antibiotics.

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• Physical agents• Mechanical removal methods• Chemical agents

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1. With minimal side effects2. Therapeutic dose: the amount of drug required for treatment or the

desired effect. 3. Broad spectrum activity: against a wide variety of pathogens or do

not know the specific bacteria that want to target.4. Chemotherapeutic agents can be synthetic or semi-synthetic.

Characteristics of antimicrobial drugs:

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Dilution Susceptibility test• Each test tube containing different concentrations of drug - MIC: minimum inhibitory concentration - MLC: minimum lethal concentration

Low [drug] high [drug]

+ + + + + + + - - -

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Image:http://www.biotopics.co.uk/microbes/penici.html

Disk diffusion test• Kirby-Bauer method• Drug diffuses from disk into agar, establishing concentration

gradient• Measure the diameter of clear zone (no growth) around disks -

> determine MIC and MLC

Large clear zone = sensitive

No or small clear zone = resistant

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Disk diffusion test

Image:http://www.biotopics.co.uk/microbes/penici.html

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How different types of antibiotics affect cell functions

Folic acid synthesis inhibitors

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Folic acid synthesis inhibitors

Sulfanilamide:

- Competitive inhibitor of PABA

- [PABA] ↑= rate of folic acid biosynthesis ↓

Image: Fdardel, 2011

DNA gyrase inhibitors

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DNA gyrase inhibitors

Quinolones:

- inhibit bacterial DNA gyrase

- effective against G- urinary tract infections and respiratory infections

- (eg. Bacillus anthracis)

- [PABA] ↑= rate of folic acid biosynthesis ↓tim

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Image: Drug Reference - Encyclopedia

Ciprofloxacin

DNA gyrase: the enzyme that introduces negative supercoils into DNA

RNA synthesis inhibitors

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RNA synthesis inhibitors

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Rifamycin/ Rifampin:

- block transcription by binding RNA polymerase

- Not selectively toxic → Prokaryotes and eukaryotes synthesize nucleic acids in pretty much the same way

- [PABA] ↑= rate of folic acid biosynthesis ↓

Cell wall synthesis inhibitors

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- Ampicillin is protected from lactamases by co-treatment with clavulanic acid

WHY: ß-lactamases have higher binding affinity for clavulanic acid than ampicillin

Image: Dengler et al. BMC Microbiology 2011 11:16   doi:10.1186/1471-2180-11-16

Cell wall synthesis inhibitors

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Image: Insilico Genomics Lab Technologies. http://insilicogenomics.in/penicillin.asp

It breaks ß-lactam rings: antibiotic resistance - G+ cells: ß-lactamases are located on outside surface

Activity is blocked by binding to transpeptidases

G- cells: ß-lactamases are in periplasmic space

(transpeptidase)

G + bacteria are more susceptible to ß-lactam antibiotics!!

blocks transpeptidization

blocks dephosphorylation of bactoprenol phosphate

blocks D-Ala peptidization

Cell wall synthesis inhibitors

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Image: Dengler et al. BMC Microbiology 2011 11:16   doi:10.1186/1471-2180-11-16

Protein synthesis inhibitors

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Protein synthesis inhibitors

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Aminoglycosides: - Binding to the small subunit

ribosome- effective against G- cells

Macrolides:- Binding to the large

subunit ribosome

Tetracyclines:- First broad-spectrum antibiotics- Blocking tRNA attachment to

ribosome- effective against G- and G+

cells

Cytoplasmic membrane inhibitors

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Cytoplasmic membrane inhibitors

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Daptomycin:

- Cyclic lipopeptide

- Makes pore on cytoplasmic membrane

- Resistance from changes in cell membrane structure

- Primarily targets G+ cells (G- cells have extra outer membrane: protection)

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Anti-fungal Drugs• Fungal infections are difficult to treat - host and pathogen have biological similarity → drug can harm host at the same time• Target against chitin (fungal cell wall) mostly - animals (host) don’t have chitin• Nystatin: first discovered antifungal antibiotic in 1949 by Hazen and

Brown• Superficial mycoses - Infections of outer layers of skin - Treatment (drugs): Miconazole, Nystatin, and Griseofulvin - Minimizes toxic systemic side effects (e.g. liver damage)

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Antiviral Drugs

• Many drugs are still in development stage• Mainly target against either RNA or DNA synthesis of viral

pathogen - Structural analogs of purine or pyrimidine bases - difficulties: viruses use metabolic machinery of the host• Protease inhibitors: against virus-specific enzymes• Interferons: stimulate production of host anti-viral proteins