antibiotics and resistance

antimicrobial agents

disinfectants
antiseptics
antibiotics

disinfectants

applied to inanimate objects - floors, tables, walls

VERY TOXIC

antiseptics

sufficiently non-toxic to be applied to living tissues

antibiotics

produced by bacteria and fungi that are exploited by humans

delivered topically and internally in a non-toxic way

most effective therapeutic agent against bacterial infections

agar plate and antibiotics

spread bacteria and let it grow

paper discs impregnated with antibiotics

KIRBY BAUER DISK INFUSION

• left - more effective, antibiotic kills

• right - ZONE OF CLEARANCE - less effective, bacteria is strong

things antibiotics can help with

availability enables chemotherapy (immunosuppressive drugs), organ transplantation, invasive surgeries, treatment of infants

problems with antibiotics

• diminished interest from pharma companies to develop new antibiotics

• bacterial resistance to antibiotics ALWAYS happen

  • unprofitable - begins in academic lab to mouse trials to a company to $1 billion trials BUT most fail before the market

deployment and resistance

1930-2005 timeline

1930-1960 —> golden age, LOTS of antibiotics discovered

1970+ —> very little new antibiotics discovered

misuse leading to antibiotic resistance

• empiric use

• increased use of broad-spectrum agents

• pediatric use for viral infections

• patients who don’t complete full antibiotic course

  • animal feeds

empiric use

blinded, use same antibiotic that you treated another patient with due to similar symptoms

• could have no idea what the actual infection is

increased use of broad spectrum agents

need to use more narrow spectrum agents if possible

pediatric use for viral infections

• don’t test for bacteria and end up being viral so antibiotics are useless here

  • ex. don’t test for strep and assume but is actually viral

patients who don’t complete full antibiotic course

• once/twice a day for two weeks

  • if stop taking once feel better, bacteria could still be present

animal feeds

• used in animals when they’re sick to prevent infection

• some antibiotics make animals gain weight —> MEATIER

• don’t want to use same antibiotic on humans and animals because once eaten, humans will possess same organisms with resistance

ON AVERAGE, humans consume 1 chicken/2 weeks

“good” antibiotic resistance will spread

measuring antibiotic activity

how effective antibiotic is in lab

MINIMUM INHIBITORY CONCENTRATION

• series of tubes, inoculate them all the same

• add increasing concentrations of antibiotics

• LOW CONC - grow fine

• HIGH CONC - doesn’t grow

• MIC - lowest conc where they can’t grow (low is good because means no resistance)

STRIPS

• agar plate with antibiotic strips, each strip contains a diff antibiotic

• certain threshold for resistance

how do antibiotics work

Target Bacterial Components

1. cell wall synthesis (beta lactams, vancomycin)

2. protein synthesis (linezolid, tetracyclines, macrolides, aminoglycosides)

3. DNA/RNA synthesis (floroquinoiones, Ri famycines)

4. folate synthesis - need folate from diet as a vitamin (Trimethoprimin, Sulfonamides)

5. cell membrane alteration (daptomycin)

TARGETS not present in eukaryotic cells

beta lactam antibiotics

• ex. penicillin

• all have a beta-lactam ring

• target cell wall

  • bind to penicillin binding proteins

    • transpeptidases - build cell wall

    • if inhibited, no peptide cross links, weak cell wall, cell death

beta lactam antibiotic resistance

• make beta lactamase - destroys ring structure (antibiotic)

Methicillin

• chemically modified penicillin that can’t be cleared by beta lactamases

• gene called mec makes a different penicillin binding protein (PBP2a)

  • doesn’t bind methicillin or other beta lactams

  • MRSA - methicillin resistant Staph aures with mec genes

vancomycin antibiotic

• old antibiotic

• inhibits cell wall synthesis in Gram+

• drug of last resort - MRSA (infection with not much left)

• binds peptide linkages at terminal D-Ala-D-Ala residues and inhibits transpeptidation

vancomycin resistance

van genes - change to D-Ala-D-Lac so can’t bind

• there is also van-resistant Staph

antibiotic resistance selection

paradox - more use of antibiotics actively selects for antibiotic resistant bacteria

antibiotic resistant strategies

• prevention of antibiotic entry - gram- outer membrane and mycobacteria cell envelope

• antibiotic modification - B-lactamase

• Efflux of antibiotics - pump out antibiotic

• alteration of target - PBPs, ribosome modification

• bypassing antibiotic action - use environmental folic acid

resistance gene

• addition to selecting for random mutations, can also be genetically encoded

  • mec, B lactamase, efflux pumps

• high levels of resistance

• classic evolution

• encoded on mobile genetic elements (ex. plasmids) allowing for HGT (superbugs)

  • eventually can’t be treated with anything

horizontal gene transfer

rather than alter gene function through mutations, new genes are acquired from another source

TRANSFORMATION - takes up DNA from environment

TRANSDUCTION - more specialized, viruses infect bacteria (bacteriophages), phages make their own genome and moves (sometimes packages wrong DNA), used in lab

CONJUGATION - plasmid encodes genes to move it, move from one bacteria to another, extremely efficient in LABS

eskape pathogen

nosocomial

• associated with healthcare system (in senior homes/hospitals)

• infect people with some underlying morbidity

ENTERCOCCUS FAECIUM (+)

STAPHYLOCOCCUS AUREUS (+)

KLEBSIELLA PNEUMONIAE (-)

ACINEIDBACTER BAUMANNIL (-)

PSEUDOMONAS AERUGINOSA (-)

ENTEROBACTER SPECIES (-)

Klebsiella Pneumoniae

• gram negative bacterium

• nosocomial pneumoniae

  • gooey capsule resistant to multiple antibiotics

• first document source of NOM-1

  • New Dehli 1 Metallo-becta-lactamases

  • Carbapenemase - destroys carbapenemes which are important beta-lactan antibiotics

  • Beta-lactamase resistant

  • NDM-1 widespread in Gram-

    • CRE (carbapenem resistant enterobacteriaceae)

      • global problem around the world

clostridia

• Gram positive

• rod-shaped

• form endospores (resistant to heat, boiling, UV light)

• strict anaerobes, vegetative cells killed by O2, spores survive

• generally found in soil and intestinal tracts of animals

• can cause life threatening diseases mediated by exotoxins

HUMAN PATHOGENS

• Clostridium Tetani

• Clostridium Botolinum

• Clostridium Perfringens

  • Clostridium Difficile

clostridium tetani

tetanus

clostridium botolinum

botulism (food-borne)

clostridium perfringens

food-borne illness and gas gangrene

clostridium difficile

exist as:

• asymptomatic carrier state in large intestine

• cause of mild-moderate diarrhea

• cause of life-threatening pseudomembranous colitis

• nosocomial - in nursing homes and hospital environments

ENDOSPORES - difficult to eradicate from environment, cultured from floor, bed pans, toilets, hands —> clothing

FECAL-ORAL ROUTE - mode of transmission

pseudo membrane colitis

pseudo membrane colitis

• antibiotic associated diarrhea, caused by C. diff

• inflammatory condition of large intestine

• RISK FACTOR - recently received antimicrobial agent

  • diarrhea, abdominal pain, fever, nausea, dehydration (1-2 days/week after antibiotic discontinued)

• endoscopy shows lesions - lesions enlarge to show inflamed mucosa + can be peeled off

  • PSEUDO MEMBRANE

• antibiotics cure infections, also kill normal microiota

• suppression of normal microbiota + persistence of C.diff endospores

• after antibiotic is stopped, spores germinate and overgrowth of C.diff occurs with production of toxins

• NOT INVASIVE - exotoxins cause damage and inflammation to lining of large intestine

• A-B TOXINS

A-B TOXINS

• produced by C.diff —> large clostudial cytotoxins

TWO DOMAINS

• A —> Active Domain Pxic, enzymatic activity —> INACTIVATES key regulatory proteins of host cells

  • B —> binding domains, uptake of host cell

C. diff treatment and diagnosis

• history (antibiotic use), symptoms and lab tests for C. diff

• endoscopy and toxin detection assays

• discontinue inciting antibiotic

• fluids

• antibiotics more specific more C.diff —> oral vancomycin / IV membrane (good for anaerobes)

  • avoid antidiarrheal agents - would cause decreased toxin clearance

fecal microbiota transplantation

new england trial

three groups

1. initial vancomucin and bowel lavage + donor infusion

2. standard vancomycin - 30%

3. standard vancomycin + bowel lavage - 23.1%

STUDY STOPPED - inappropriate to keep patients on standard treatment when donor was so effective