BETA LACTAM ANTIBIOTICS PENICILLINS & CEPHALOSPORINESEMRE, THE BARBARIAN YEDITEPE (SEVEN HILL? ) FALL 2010

PATHOGENIC BACTERIAFIRST, A BRIEF INTRODUCTION AND SUMMARY OF PATHOGENIC BACTERIA

Eukaryotic and Prokaryotic CellsProkaryotes (GR: Before nucleus)y No membrane around

Eukaryotes (GR: True nucleus)y Nuclear membrane -

DNA NO NUCLEUS. y Smaller than eukaryotes (approx. 1.0 m in diameter) y Bacteria and archaea. y Simple and small

NUCLEUS y 10-100 m y Algae, protozoa, fungi, animals, plants y Large and more complex than prokaryotes

Prokaryote cell

Another picture of prokaryotic cell

Eukaryote cell

Another picture of eukaryotic cell

comparisonWhat major difference? Eukaryotes contain nuclei, visible with light microscopy. Prokaryotes lack nuclei. What else? Prokaryotic ribosomes are 70S and composed of 30S and 50S subunits. Eukaryotic ribosomes are 80S with 60S and 40S subunits.

Morphology of prokaryotic cells

PATHOGENIC BACTERIAy BOTH WONDERFUL, POWERFUL AND HORRIBLE

CREATURES THAT SELF-REPLICATE AND SURVIVE IN THE HARSH AND HOSTILE ENVIRONMENT OF THE HUMAN BODY.

The major groups of medically important bacteriaThis illustration is a practical representation of the principal groups of pathogenic bacteria. It is meant to be a study aid, not a taxonomic or phylogenetic tree.

USUAL SUSPECTS & KOCHy Although the vast majority of bacteria are harmless or

beneficial, quite a few bacteria are pathogenic.y LETS REMEMBER SOME USUAL SUSPECTS .

y TUBERCULOSIS: Mycobacterium tuberculosis kills about 2

million people a year, mostly in sub-Saharan Africa. y PNEUMONIA: Streptococcus and Pseudomonas y FOODBORNE INFECTIONS: Shigella, Campylobacter and Salmonella.Pathogenic bacteria also cause infections such as tetanus, typhoid fever, diphtheria, syphilis and leprosy. Koch's postulates are criteria designed to establish a causal relationship between a causative microbe and a disease.

KOCHS POSTULATESy Koch's postulates are four criteria

designed to establish a causal relationship between a causative microbe and a disease. The postulates were formulated by Robert Koch and Friedrich Loeffler in 1884 and refined and published by Koch in 1890. Koch applied the postulates to establish the etiology of anthrax and tuberculosis, but they have been generalized to other diseases.

KOCHS FOUR POSTULATES1.

2. 3. 4.

The microorganism must be found in abundance in all organisms suffering from the disease, but should not be found in healthy animals. The microorganism must be isolated from a diseased organism and grown in pure culture. The cultured microorganism should cause disease when introduced into a healthy organism. The microorganism must be reisolated from the inoculated, diseased experimental host and identified as being identical to the original specific causative agent.

Pathogenic bacteria: conditional or intracellulary CONDITIONALLY PATHOGENIC:y PART OF THE HUMAN FLORA y PATHOGENIC ONLY UNDER CERTAIN CONDITIONS:

WOUND OR IMMUNODEFICIENCY y Staphylococcus or Streptococcus SPECIES y Pseudomonas aeruginosa, Burkholderia cenocepacia, and Mycobacterium avium, are opportunistic pathogens and cause disease mainly in people suffering from immunosuppression or cystic fibrosis.y INTRACELLULAR

INTRACELLULAR PATHOGENSy Facultative intracellular parasites are capable of living and

reproducing either inside or outside cells.y Bacterial examples include Neisseria meningitidis, Francisella

tularensis and Listeria monocytogenes. Other examples include Brucella, Legionella, Mycobacterium, and Yersinia. y A fungal example is Histoplasma capsulatum.y Obligate intracellular parasites cannot reproduce outside their

host cell, meaning that the parasite's reproduction is entirely reliant on intracellular resources.

OBLIGATEy Viruses y Certain bacteria, including:y Chlamydia, and closely

y Certain protozoa,

including:y Plasmodia species y Leishmania spp. y Toxoplasma gondii y Trypanosoma cruzi

related species.[3] y Rickettsia y Coxiella y Certain species of Mycobacterium such as Mycobacterium leprae

The mitochondria in eukaryotic cells may also have originally been such parasites, but ended up forming a mutualistic relationship (endosymbiotic theory). Study of obligate pathogens is difficult because they cannot usually be reproduced outside the host. However, in 2009 scientists reported a technique allowing the Q-fever pathogen Coxiella burnetii to grow in an axenic culture and suggested the technique may be useful for study of other pathogens.

ENDOSYMBIOTIC THEORYThe endosymbiotic theory concerns the origins of mitochondria and plastids (e.g. chloroplasts), which are organelles of eukaryotic cells. According to this theory, these organelles originated as separate prokaryotic organisms that were taken inside the cell as endosymbionts. Mitochondria developed from proteobacteria (in particular, Rickettsiales or close relatives) and chloroplasts from cyanobacteria.

EXAMPLES OF NORMAL BACTERIAL FLORA (NORMAL MICROBIOTA)

THE HUMAN BODY CONTAINS A VAST NUMBER OF PATHOGEN ORGANISMS THAT MAKE UP THE NORMAL HUMAN FLORA. We must know what microorganisms live in what parts of our body to decide whether there is an infection or a colonization.

Descriptiony Contamination mere presence of microbes in or on the body. Contamination:y Food, drink or air: External contaminants. y Wounds, biting arthropods (bugs), sexual intercourse.

y Colonization microbial contaminants remain where they first Colonization:

contacted the body without causing any harm (Staphylococcus epidermidis on the skin; pathogens in mucous membranes) and they become part of the RESIDENT MICROBIOTA or got rid of and become TRANSIENT MICROBIOTA. y Infection successful invasion of the body by a pathogen. The Infection: pathogen that entered the body through contamination overcame the bodys defenses.

Resident microbiota I

Ref: Chapther 14: Infection, Infectious Diseases and Epidemiology. Microbiology; Alternate Edition with Diseases by Body System. Robert W. Bauman, Ph.D. Pearson Benjamin Cummings. 2006. pp 408-409

Resident microbiota II

How normal microbiota becomes opportunistic pathogensy IMMUNE SUPPRESSIONy Disease (AIDS, renal insufficiency, hepatitis), malnutrition,

emotional stress, physical stress, extremes of age (very young or very old), radiation, chemotherapy, immunosuppressive drugsy CHANGES IN THE NORMAL MICROBIOTAy Use of antimicrobials can lead to formation of Candida albicans

in the vagina. y Hormonal changes, stress, changes in diet y Exposure to overwhelming number of pathogensy INTRODUCTION OF A MEMBER OF THE NORMAL

MICROBIOTA INTO AN UNUSUAL SITE IN THE BODYy Ex. Rectal-vaginal transmission of E coli, entering urethra

Let us remember Kochs postulatesy It is critical that all the postulates are satisfied in this order.

The suspected pathogen must be present in every case of the disease. 2. The pathogen must be isolated and grown in pure culture. 3. The cultured pathogen must cause the disease when it is inoculated into a healthy, susceptible experimental host. 4. The same pathogen must be reisolated from the diseased experimental host.1.

Exception to Kochs postulatesy Some pathogens can not be cultured in the laboratory:y Mycobacterium leprae

y Some diseases are cuased by a combination of pathogens OR

a combination of pathogen and other cofactors (environmental or genetic):y Liver cancer: when both hepatitis B and hepatitis D

y Ethical considerations prevent applying to Kochs postulates

to diseases and pathogens that occur in HUMANS ONLY!y AIDS: observations of fetuses of infected mothers or of

healthcare workers who were infected accidentally

Difficulties against Kochs postulatesy Single cause not possible:y Pneumonia, meningitis, hepatitis

y Ignoring some pathogensy Helicobacter pylori and gastric ulcers

Virulence factors of Infectious Agentsy Pathogenicity: the ability of a mo to cause disease. y Virulence: the degree of pathogenicity. y Virulence: the relative ability of a pathogen to infect a host

and cause disease. y Virulence factors: traits of pathogens that enable them to enter a host, adhere to host cells, gain access to nutrients, escape detection or removal of the host cell.

Virulence continuumy More virulenty Francisella tularensis (rabbit fever) y Yersinia pestis (plague) y Bordetella pertussis (whooping cough) o Pseudomonas aeruginosa (infections of burns) Clostridium difficile (antibiotic induced / pseudomembraneuse collitis) Candida albicans (vaginitis, thrush) Lactobacilli (diphtheroidis) Less virulent

The Big Four typical bacteriay The four categories presented in Figure are fairly evenly y y y y y

represented in the normal flora of: the mouth, the pharynx, and the large intestine; major pathogens, however, are represented in fewer numbers. The Gram-positive cocci and the Gram-negative rods are the most common agents of infection, followed by the Gramnegative cocci and the Gram-positive rods.

Diseases caused by selected Gram (+) BacteriaGenera Clostridium Characteristics Obligate anaerobic rods (endospore formers) Diseases Tetanus Botulism Gangrene Severe diarrhea (pseudomembraneouse collitis) Dental caries Dental caries

Epulopiscium Veillonella

Giant rods Part of oral biofilm on human teeth; stain like Gram (-) / PINK No cell wall; pleomorphic, smallest free-living cells, stain like Gram (-) / PINK Facultative anaerobic rods; endospore formers Contaminates of dairy products

Mycoplasma

Pneumonia Urinary tract infections Anthrax Listeriosis

Bacillus Listeria

Diseases caused by selected Gram (+) BacteriaGenera Lactobacillus Streptococcus Staphylococcus Corynebacterium Characteristics Produce yogurt, buttermilk, pickles, sauerkraut Cocci in chains Cocci in clusters Snapping division; metachromatic granules in cytoplasm Waxy cell walls (mycolic acid) Filaments Filaments; degrade pollutants Produce antibiotics Diseases Rare blood infections Strep throat, scarlet fever, and others Bacteremia, food poisoning, and others Diphtheria

Mycobacterium Actinomyces Nocardia Streptomyces

Tuberculosis and meningitis Actinomycosis Lesions Rare sinus infections

Diseases caused by selected Gram (-) BacteriaGenera Rickettsia Brucella Ehrlichia Neisseria Burkholderia Legionella Coxiella Pseudomonas Intracellular pathogens Intracellular pathogens Aerobes that catabolize carbohydrates via EntnerDoudoroff and pentose phosphate way Characteristics Intracellular pathogens Coccobacilli Live inside white blood cells Diplococci Diseases Typhus Rocky Mountain spotted fever Brucellosis Leukopenia Gonorrhea and meningitis Lung infection of cystic fibrosis patients Legionnaires disease Q fever Urinary tract infections, external otitis

Diseases caused by selected Gram (-) BacteriaGenera Campylobacter Helicobacter Treponema Borrelia Bacteroides Escherichia Salmonella Proteus Shigella Yersinia Klebsiella Vibrio Haemophilus Vibrios (Vibrionaceae) Cocci or straight rods (Pasteurellaceae) Characteristics Curved rods Spirals Spirochetes Spirochetes Anaerobes that live in animal colons Diseases Gastroenteritis Gastric ulcers Syphilis Lyme disease Abdominal infections

Straight rods Gastroenteritis (Enterobacteriaceae) Facultative Enteritis Anaerobes Urinary tract infections Shigellosis Plague Pneumonia Cholera Meningitis in children

ANTIBIOTICSWHAT ARE THEY? HOW DO THEY WORK?

ANTIBIOTICS secondary metabolitesy MEMBERS of extremely diverse group of metabolites known y y y y y

as secondary metabolites. Secondary metabolites are complex organic molecules that are not essential for normal cell growth. Tetracyline: 72 separate enzymatic steps (Streptomyces) Erythromycine: 25 different chemical reactions (Streptomyces). Therefore, antibiotics are expensive to bacteria. Also: in natural soil, antimicrobials are not found at concentrations inhibitory to neighbouring cells.

Why are there antibotics for m.o.?y Leftovers of metabolic pathways that were once beneficial? y Truly required by the host cell to defend itself? y These hypotheses are not proven, but y Antibiotics are very useful for the mankind to defend against

infections.

Sources of some common antibioticsMICROORGANISM ANTIMICROBIAL FUNGI Penicillum chrysogenum Penicillium griseofulvin Cephalosporium spp. BACTERIA Bacillus licheniformis Bacillus polymyxa Streptomyces griseus Streptomyces fradiae Bacitracin Polymyxin Streptomycin Neomycin Penicillin Griseofulvin Cephalothin MICROORGANISM ANTIMICROBIAL BACTERIA CONTINUED Streptomyces orientalis Streptomyces venezuelae Streptomyces erythraeus Streptomyces nodosus Streptomyces avermitilis Streptomyces aureofaciens Vancomycine Chloramphenicol Erythromycin Amphotericin B Ivermectin Tetracycline

Micromonospora purpurea Gentamicin

Streptomyces mediterranei Rifampin

Antibiotics Selectiony Culture and sensitivity testing prior to antibiotic prescribing. y Empirical antibiotic therapy should be based on:y Knowledge of likely pathogens for the site of infection y Anamnesis and patient history: recent hospitalizations, work-

related exposure, travel, pets, etc. y Local susceptibility

THREE MAIN WAYS TO ATTACK PATHOGENIC BACTERIAy KNOW THY ENEMY AND KNOW YOURSELF; IN A

HUNDRED BATTLES YOU WILL NEVER BE IN PERIL. THE ART OF WAR, SUN TZU

1. 2. 3.

THE BACTERIAL CELL ENVELOPEy A UNIQUE STRUCTURE NOT PRESENT IN HUMANS

BIOSYNTHETIC PROCESSES WITHIN THE BACTERIAy BACTERIAL PROTEIN PRODUCTION

BACTERIAL REPLICATION (DNA)

Antibiotic Mechanisms of Action

Mechanisms of action

Beta lactams attack cell wall what is it?y Most bacteria have a cell wall,

y y

y y

composed of PEPTIDOGLYCAN. Peptidoglycan is a complex polysaccharide. Peptidoglcan is composed of two regularly alternating sugars: NAM: N-Acetylmuramic Acid NAG: N-Acetylglucosamine

Structure of peptidoglycan

Comparison of cell walls Gram (+) & Gram (-)

What is the effect of Lipid A on humans? Lipid A causes shock, blood clotting and fever in humans!

CELL ENVELOPEGRAM POSITIVEy CYTOPLASMIC MEMBRANE

GRAM NEGATIVEy CYTOPLASMIC MEMBRANE

SURROUNDED BY TOUGH AND RIGID CELL WALL y GRAM STAINING: BLUE (PURPLE-positive-P)

SURROUNDED BY THE OUTER MEMBRANE (THIN, LIPID MEMBRANE) y GRAM STAINING: PINK (negative-N)

CYTOPLASMIC MEMBRANEy PREVENTS IONS FROM FLOWING INTO OR OUT OF y y y y

THE CELL ITSELF. MAINTAINS THE CYTOPLASM AND BACTERIAL COMPONENTS IN A DEFINED SPACE. GRAM (+) / CELL WALL: PROTECTS IT FROM MECHANICAL AND OSMOTIC STRESSES. GRAM (-) / MEMBRANE: CONTAINS PORINS (ION CHANNELS) HUMAN CELLS: DO NOT CONTAIN A CELL WALL.

STRUCTURE OF THE CELL WALLy PEPTIDOGLYCAN (LONG SUGAR POLYMERS)y N-ACETYL-GLUCOSAMINE + N-ACETYL-MURAMIC

ACIDy + PEPTIDE SIDE CHAINS y CROSS-LINKED TOGETHERy JUST AS CROSS-LINKING OF METAL LOOPS

STRENGTHENED THE CHAIN MAIL ARMOR USED BY MEDIEVAL KNIGHTS.y CROSS-LINKING IS MEDIATED BY PENICILLIN-

BINDING PROTEINS (PBPs): BACTERIAL ENZYMES

Schematic of typical GRAM-POSITIVE cell wall showing arrangement of N-Acetylglucosamine GRAMand N-Acetylmuramic acid; Teichoic acids not shown.

Schematic of typical GRAM-NEGATIVE cell wall showing arrangement of N-Acetylglucosamine GRAMand N-Acetylmuramic acid and the outer membrane containing lipopolysaccharide.

A general view of all antibiotics

GRAM POSITIVE & NEGATIVE?A Gram stain of mixed Staphylococcus aureus (Gram positive cocci, purple) and Escherichia coli (Gram negative bacilli, pink). (Image:Y Tambe, Wikimedia Commons)

ANTIBACTERIAL AGENTS: BETA LACTAMSPENICILLINS & CEPHALOSPORINES: ATTACKING THE CELL MEMBRANE IN FIGURE, THE STRUCTURE OF THE BETA LACTAM RING IS SHOWN.

-Lactam Antibioticsy The essential core of penicillin is a four-member ring called a

-lactam ring. y Modifications led to penicillins, cephalosporins, carbapenems, and monobactams. y -lactam antibiotics: inhibitors of penicillin-binding proteins (PBPs) that normally assemble the peptidoglycan layer surrounding most bacteria.

-Lactams: an important class of antibacterial compounds.y Cyclic amides with four atoms in the ring and this ring is

more accurately referred to as an azetidinone ring. Examples of -lactams include y penicillins (natural and synthetic), y cephalosporins (natural and synthetic), cephems (also known as carbacephems), cephams, penems, penams, carbapenems, cephamycins, 1-oxacephems, clavulanic acids (also known as clavams), nocardicins, trinems (also known as tribactams) and monobactams.

-Lactams: an important class of antibacterial compounds.

Mechanism of action of -lactam antibioticsA. Normally, a new subunit of Nacetylmuramic acid (NAMA and NNAMA) acetylglucosamine (NAGA disaccharide NAGA) with an attached peptide side chain is linked to an existing peptidoglycan polymer. This may occur by covalent attachment of a glycine (G) bridge from one peptide side chain to another through the enzymatic action of a penicillin-binding protein (PBP). B. In the presence of a lactam antibiotic, this process is disrupted. The -lactam antibiotic binds the PBP and prevents it from cross-linking the glycine bridge to the peptide side chain, thus blocking incorporation of the disaccharide subunit into the existing peptidoglycan polymer.

How does -Lactam ring do that?y The -lactam ring mimics the D-alanyl-D-alanine portion of

the peptide side chain that is normally bound by PBPs. PBPs thus interact with the -lactam ring and are not available for synthesis of new peptidoglycan. y The disruption of the peptidoglycan layer leads to lysis of the bacterium.

Mechanism of penicillin-binding protein (PBP) inhibition by -lactam antibiotics.A. PBPs recognize and catalyze the peptide bond between two alanine subunits of the peptidoglycan peptide side chain. B.The -lactam ring mimics this peptide bond. Thus, the PBPs attempt to catalyze the -lactam ring, resulting in inactivation of the PBPs.

RESISTANCE TO BETA LACTAMSINTRINSICy Pseudomonas aeruginosa y the porins in its outer

ACQUIREDy Pseudomonas aeruginosa y The carbapenem

membrane strains do not allow passage of ampicillin to the periplasmic space.

imipenem gains access to imipenem the PBPs of this organism by passing through a specific protein channel found in the outer membrane. spontaneous mutations

RESISTANCE TO BETA LACTAMSINTRINSICy LL STRAINS ARE

ACQUIREDy SOME STRAINS ARE

RESISTANT.

RESISTANT.

THE SIX PS (SIX POTENTIAL PITFALLS)1. 2. 3. 4. 5. 6.

Penetration Porins Pumps - efflux pumps Penicillinases (really -lactamases, but that does not start with P!) PBPs Peptidoglycan is absent

Six P's by which the action of -lactams may be blocked.

Six P's by which the action of -lactams may be blocked.1. 2. 3. 4. 5.

6.

Penetration Porins Pumps Penicillinases ( lactamases) Penicillinbinding proteins (PBPs) Peptidoglycan.

1/6. PENETRATION-lactams penetrate poorly into the intracellular compartment of human cells, so bacteria that reside in this compartment are not exposed to them. y A -lactam antibiotic cannot kill a bacterium if it cannot get to it! y Remember: Facultative and Obligate microorganisms.y

2/6. PORINSy If a -lactam antibiotic does reach the bacterium, it must gain

access to its targets, the PBPs. y In Gram-positive bacteria, this is not difficult because the PBPs and the peptidoglycan layer are relatively exposed, but Gramin Gram-negative bacteria they are surrounded by the protective outer membrane. y -Lactams must breach this membrane by diffusing through porins, which are protein channels in the outer membrane. y Many Gram-negative bacteria have porins that do not allow passage of certain -lactams to the periplasmic space.

3/6. PUMPS (EFFLUX PUMPS)y Pumpssome bacteria produce efflux pumps, which are

protein complexes that transport antibiotics, which have entered the periplasmic space, back out to the environment. y These pumps prevent antibiotics from accumulating within the periplasm to concentrations sufficient for antibacterial activity.

4/6. PENICILLINASES ( -LACTAMASES)y Penicillinases: really -lactamases, but that does not start

with P!) y Many bacteria, both Gram-positive and Gram-negative, make -lactamases, enzymes that degrade -lactams before they reach the PBPs.

5/6. PBPsy Some bacteria produce PBPs that do not bind -lactams with

high affinity. y In these bacteria, -lactams reach their targets, the PBPs, but cannot inactivate them.

6/6. PEPTIDOGLYCANy Peptidoglycan is absentthere are a few bacteria that do not

make peptidoglycan and that therefore are not affected by lactams.

EFFECTIVE -LACTAMASE INHIBITORS* ARE EFFECTIVE IF:y THEY CAN SUCCESSFULLY NAVIGATE AROUND

POTENTIAL PITFALLS: 6PS:1. 2. 3. 4. 5. 6.

PENETRATION PORINS PUMPS - EFFLUX PUMPS PENICILLINASES (really -lactamases, but that does not start with P!) PBPs PEPTIDOGLYCAN IS ABSENT

* = -LACTAM AGENTS (= -LACTAMS)

-LACTAMASEs come in many flavors!y the -lactamase of Staphylococcus aureus is relatively specific

for some of the penicillins. y the extended-spectrum -lactamases made by some strains of Escherichia coli and Klebsiella spp. degrade nearly all penicillins, cephalosporins, penicillins cephalosporins and monobactams monobactams.

PENICILLINSPictures show Penicillium species, from which penicillins were derived: Penicillium notatum, P chrysogenum,

Discovery of penicillinsy Alexander Fleming: 2 week vacation y Incubation left at room temperature

instead of 37C

y to slow the growth rate of the bacteria

y Penicillium grows at room

temperature but not 37C.

y VACATIONS MAKE TRUTHFULLY

PRODUCTIVE AT WORK!

y GO ON HOLIDAY

TO MAKE A DISCOVERY.

a thiazolidine ring attached to a -lactam ring that is itself modified by a variable side chain (R in Fig.).6-APA: 6-Aminopenisilanic acid = -lactam ring + THIAZOLIDINE RINGTHE STRUCTURE OF PENICILLINS

CHEMICAL STRUCTURE - FUNCTIONSy THIAZOLIDINE -LACTAM : antibacterial activity y SIDE CHAIN: extra pharmacologic activity:

Types (per side chain)y Natural Penicillins y Antistaphylococcal penicillins y Aminopenicillins y Extended-spectrum penicillins

+y Penicillins in combination with

-lactamase inhibitors

TR Brands Category Natural penicillins Parenteral Agents Penicillin G TR Brands Penadur , Deposilin Oral Agents Penicillin V Pen-Os (Ospen)*, Cliacil **,

Antistaphylococcal penicillins Aminopenicillins

Nafcillin, oxacillin, (methicillin) Ampicillin

Dicloxacillin

Amoxicillin, ampicillin Alfasid, Duocid Amoxicillinclavulanate Carbenicillin

Alfoxil, Largopen*** Alfasilin, Silina**** Augmentin, Amoklavin, Croxilex

Aminopenicillin + lactamase inhibitor Extended-spectrum penicillins Extended-spectrum penicillin + lactamase inhibitor

Ampicillinsulbactam Piperacillin, ticarcillin Piperacillintazobactam, ticarcillinclavulanate

Geopen

Tizacin, Timentin

* Penisilin V benzatin, **penisilin V potasyum, ***amoxicillin, **** ampicillin

SCHEMATIC TO UNDERSTAND EFFICACYy WALK SIGN: ACTIVE

y STOP SIGN: NOT ACTIVE

y CAUTION SIGN: SOMETIMES ACTIVE

SCHEMATICS TO DERIVE GENERAL EFFICACY / examplesy TO understand clinically what to do with what bacteria; y y y y y

lets divide bacteria into four major groups: Gram-positive, Gram-negative, anaerobes, atypical. Gram-positive: GO! Use the antibiotic to treat infection. (ACTIVE) Gram-negative: STOP! Dont use the antibiotic. (NOT ACTIVE). Anaerobes: PROCEED WITH CAUTION! (SOMETIMES ACTIVE) Atypical: PROCEED WITH CAUTION! (SOMETIMES ACTIVE)

Let us remember the Gram negative and the Gram positive. positive

NATURAL PENICILLINSWhich is oral and which is parenteral?

RESISTANCE TO NATURAL PENICILLINSy Some are intrinsically resistant and some have acquired

resistance: Six Ps again: What were the six Ps? 1. PENETRATION 2. PORINS 3. PUMPS - EFFLUX PUMPS 4. PENICILLINASES (really -lactamases, but that does not start with P!) 5. PBPs 6. PEPTIDOGLYCAN IS ABSENT

KNOW THY ENEMY: CHOOSE YOUR WEAPON

1. Penetration: Rickettsia & Legionellay Penetrationnatural penicillins, like most -lactams,

penetrate poorly into the intracellular compartment of human cells, so bacteria that for the most part reside in this compartment, such as y Rickettsia and y Legionella, y are protected from them.

2. Porinsy PorinsSome Gram-negative bacteria, such as y Escherichia coli, y Proteus mirabilis, y Salmonella enterica, and y Shigella spp., y have porins in their outer membranes that do not allow

passage of the hydrophobic natural penicillins to the periplasmic space.

3. Pumpsy Pumpssome Gram-negative bacteria, such as y Pseudomonas aeruginosa, y have efflux pumps that prevent the accumulation of

penicillins within the periplasm. Although these pumps by themselves may only cause a marginal change in susceptibility, they can work together with penicillinases and porins to have a dramatic effect.

4. Penicillinasesy Penicillinases y many bacteria, y both Gram-positive (staphylococci) and y Gram-negative (some Neisseria and Haemophilus strains, many

of the enteric species not listed in [b], and y some anaerobes, such as Bacteroides fragilis), y make penicillinases that degrade the natural penicillins.

5. PBPsy Penicillin-binding proteins (PBPs)some bacteria produce

PBPs that do not bind natural penicillins with a high affinity (e.g., some strains of y Streptococcus pneumoniae).

6. Peptidoglycany Peptidoglycansome bacteria, such as y Mycoplasma and y Chlamydia spp., y do not make peptidoglycan and therefore are not affected by

the natural penicillins.

Natural Penicillinsy can be purified directly from cultures of Penicillium mold. y R (penicillin G): hydrophobic benzene ring.

y Nearly all bacterias cell wall consist of peptidoglycan. y Active against:y some species of Gram-positive, y Gram-negative, and y anaerobic bacteria, as well as some spirochetes.

ANTIMICROBIAL ACTIVITY OF NATURAL PENICILLINSBACTERIA Gram-positive bacteria BACTERIATYPE Streptococcus pyogenes Viridans group streptococci Some Streptococcus pneumoniae Some enterococci Listeria monocytogenes Neisseria meningitidis Some Haemophilus influenzae Clostridia spp. (except C. difficile) Actinomyces israelii

Gram-negative bacteria Anaerobic bacteria

Atypical bacteria

Treponema pallidum Leptospira spp.

Take home lesson!y Even a few Gram-negative bacteria, y such as Neisseria meningitidis and some strains of Haemophilus

influenzae that do not make -lactamases, remain susceptible to penicillin.

Lets remember: Gonna need it in the upcoming slides.y WALK SIGN: ACTIVE

y STOP SIGN: NOT ACTIVE

y CAUTION SIGN: SOMETIMES ACTIVE

SCHEMATICS TO DERIVE GENERAL EFFICACY / examplesy Gram-positive: GO! Use the antibiotic to treat

infection. (ACTIVE)y Gram-negative: STOP! Dont use the antibiotic.

(NOT ACTIVE).y Anaerobes: PROCEED WITH CAUTION!

(SOMETIMES ACTIVE) y Atypical: PROCEED WITH CAUTION! (SOMETIMES ACTIVE)

ANTISTAPHYLOCOCCAL PENICILLINSPENICILLINASE-RESISTANT PENICILLINS: NAFCILLIN OXACILLIN DICLOXACILLIN FLUOXACILLIN CLOXACILLIN

R side chain of nafcillin.

Antistaphylococcal Penicillinsy BIG residues on their R side chains that prevent binding by

the staphylococcal -lactamases. y Thus: penicillinase resistant -lactam penicillins y TREATS:y Staphylococcus aureus y Staphylococcus epidermidis

y BUT CANNOT TREAT:y MRSA: Methicillin-Resistant Staphylococcus aureus y MRSE: Methicillin-Resistant Staphylococcus epidermidis

y Because they can not bind to the PBPs of MRSA and MRSE.

METHICILLIN? You did not mention that!y an antistaphylococcal penicillin that is no longer

commercially available y Representative of the entire class of antistaphylococcal penicillins in its spectrum of activity. y less effective than natural penicillins against STREPTOCOCCI and are usually not used to treat them. y less effective against ENTEROCOCCI and are usually not used to treat them.

ANTIMICROBIAL ACTIVITIY OF ANTISTAPHYLOCOCCAL PENICILLINSBACTERIA Gram-positive bacteria BACTERIATYPE Some Staphylococcus aureus Some Staphylococcus epidermidis

AMINOPENICILLINSAMPICILLIN AMOXICILLIN

R side chain of ampicillin

AMINOPENICILLINSy An additional amino group y increases their hydrophilicity y INCREASED HYDROPHILICITY allows them to pass

through the PORINS in the outer membranes of some enteric Gram-negative rods. Ex: y E. coli, P. mirabilis, S. enterica, and Shigella spp.

ANTIMICROBIAL ACTIVITY OF AMINOPENICILLINSBACTERIA Gram-positive bacteria BACTERIATYPE Streptococcus pyogenes Viridans group streptococci Some Streptococcus pneumoniae Some enterococci Listeria monocytogenes Neisseria meningitidis Some Haemophilus influenzae Some Enterobacteriaceae Clostridia spp. (except C. difficile) Actinomyces israelii

Gram-negative bacteria

Anaerobic bacteria

Atypical bacteria:Spirochets

Borrelia burgdorferi

Aminopenicillin/ -Lactamase Inhibitor Combinationsy CLAVULANATE and SULBACTAM y Compounds developed to inhibit the -lactamases of many

Gram-positive and Gram-negative bacteria y structurally similar to PENICILLIN and therefore bind lactamases, which results in the inactivation of the lactamases. y Ampicillin-sulbactam / parenteraly (sultamisilin prodrug) / oral

y amoxicillin-clavulanate / oral (parenteral?)* y broaden the antimicrobial spectrum of the aminopenicillins. y *AUGMENTIN I.V.

ANTIMICROBIAL ACTIVITY OF AMINOP. + -LACTAMASE INHIBITOR COMBOSBACTERIA Gram-positive bacteria BACTERIATYPE Some Staphylococcus aureus Streptococcus pyogenes Viridans group streptococci Some Streptococcus pneumoniae Some enterococci Listeria monocytogenes Neisseria spp. Some Haemophilus influenzae MANY Enterobacteriaceae Clostridia spp. (except C. difficile) Actinomyces israelii Bacteroides spp. Borrelia burgdorferi

Gram-negative bacteria

Anaerobic bacteria

Atypical bacteria: Spirochets

EXTENDED SPECTRUM PENICILLINSPIPERACILLIN TICARCILlIN CARBENICILLIN Even greater penetration into Gram-negative bacteria than is seen with the aminopenicillins

R side chain of piperacillin

EXTENDED SPECTRUM PENICILLINSy Even greater penetration into Gram-negative bacteria than is

seen with the aminopenicillins. y Best example: the side chain of piperacillin is polar. y increases its ability to pass through the outer membrane porins of some Gram-negative bacteria. y Piperacillin gets its name from its side chain, which contains a piperazine derivative.

Resistance and activityy more resistant to cleavage by Gram-negative -lactamases. y more active against Gram-negative bacilli, including many

strains of P. Aeruginosa. y susceptible to the -lactamases of staphylococci (unlike natural penicillins!) y modest activity against anaerobes. y piperacillin has the broadest activity.

ANTIMICROBIAL ACTIVITY OF EXTENDED SPECTRUM PENICILLINSBACTERIA Gram-positive bacteria BACTERIATYPE Streptococcus pyogenes Viridans group streptococci Some Streptococcus pneumoniae Some enterococci Neisseria meningitidis Some Haemophilus influenzae Some Enterobacteriaceae Pseudomonas aeruginosa Clostridia spp. (except C. difficile) Bacteroides spp.

Gram-negative bacteria

Anaerobic bacteria

Extended-Spectrum Penicillin/ Lactamase Inhibitor Combinationspiperacillin-tazobactam and ticarcillin-clavulanate

Ext.Spect. Penicillin/ -Lactamase Inh. Combinationsy Strategy: combine extended-spectrum penicillins with -

lactamase inhibitors. y piperacillin-tazobactam y ticarcillin-clavulanate y The -lactamase inhibitors neutralize many of the lactamases that otherwise inactivate the extended-spectrum penicillins, resulting in a marked enhancement of their activity.

Activityy activity against most aerobic Gram-positive bacteriay including many -lactamaseproducing staphylococci

y most aerobic Gram-negative bacteria y nearly all anaerobic bacteria except Clostridium difficile y piperacillin-tazobactam has a broader spectrum than

ticarcillin-clavulanate.

ANTIMICROBIAL ACTIVITY OF EXT.D-SPECT. PENICILLIN + -LACTAMASE INH. COMBOSBACTERIA Gram-positive bacteria BACTERIATYPE Some Staphylococcus aureus Streptococcus pyogenes Viridans group streptococci Some Streptococcus pneumoniae Some enterococci Listeria monocytogenes Neisseria spp. Haemophilus influenzae MOST Enterobacteriaceae Pseudomonas aeruginosa Clostridia spp. (except C. difficile) Bacteroides spp.

Gram-negative bacteria

Anaerobic bacteria

TOXICITY / ADVERSE REACTIONS OF PENICILLINSAllergy: GENERAL RULE OF THUMB: An estimated 3%10% of people are allergic to penicillins. Adverse Reactions: nausea, vomiting, and diarrhea. Also (less likely): drug fever, rash, serum sickness, interstitial nephritis, hepatotoxicity, neurologic toxicity, and hematologic abnormalities. HYPERSENSITIVITY: Urticaria, angioedema, and anaphylaxis. GOLDEN RULE: Persons allergic to one penicillin should be considered allergic to all penicillins, and cross-allergenicity may extend to other lactam antibiotics.

TOXICITY / ADVERSE REACTIONSy Allergy: GENERAL RULE OF THUMB: An estimated 3% y y

y y

10% of people are allergic to penicillins. Adverse Reactions: nausea, vomiting, and diarrhea. Also (less likely): drug fever, rash, serum sickness, interstitial nephritis, hepatotoxicity, neurologic toxicity, and hematologic abnormalities. HYPERSENSITIVITY: Urticaria, angioedema, and anaphylaxis. GOLDEN RULE: Persons allergic to one penicillin should be considered allergic to all penicillins, and cross-allergenicity may extend to other -lactam antibiotics.

SUMMARY OF ACTIVITY OF PENICILLINSy The antistaphylococcal penicillins are inactive against Gramramy

y

y y

negative bacteria. The natural penicillins have activity against N. meningitidis and some strains of H. influenzae, but few other Gram-negative influenzae bacteria. The spectrum of the aminopenicillins is expanded to include these organisms plus some enteric Gram-negative rods such as certain Gramrods, strains of E. coli, P. mirabilis, S. enterica, and Shigella spp that do not mirabilis, enterica, spp., produce -lactamases. The extended-spectrum penicillins are active against even more xtendedaeruginosa enteric Gram-negative rods and, importantly, P. aeruginosa. Finally, the addition of a -lactamase inhibitor to an extendedextended-spectrum penicillin extends this list to include most enteric Gram-negative bacilli. Gram-

MNEMONIC: REMEMBERCLASS OF PENICILLIN ANTISTAPHYLOCOCCAL PENICILLINS NATURAL PENICILLINS AMINOPENICILLINS EXTENDED SPECTRUM PENICILLINS EXTENDED-SPECTRUM PENICILLINS + LACTAMASE INHIBITOR MNEMONIC N (Nursing) NH (Nursing Home) NHFE (Nursing Home Few Elderly) NHSEP (Nursing Home Some Elderly Persons) NHMEP (Nursing Home Most Elderly Persons) COVERAGE NONE Neisseria meningitidis, some Haemophilus + few enteric rods +some enteric rods, Pseudomonas aeruginosa +most enteric rods, Pseudomonas aeruginosa

CASE STUDY 48 YEAR OLD MANA 48 YEAR OLD MAN.

48 years old male with coughy A 48 year old man comest to your pharmacy with a 6 day y y y

y

history of worsening cough productive of green sputum. He has had fever and chills. He complains of pain in the right midback with deep breathing or coughing. T=38.1C (100.5F), his respiratory rate: 24 breaths per minute, pulse: 98 beats per minute, blood pressure: 120/75 mmHg, saturation of oxygen: 96 % on room air by pulse oximetry. Auscultation of his lungs reveals rales in the right lowerposterior lung field.

What would you treat him with?y A posterior-to-anterior (PA) and lateral chest x-ray show a y y y y

right lower-lobe infiltrate. A sputum Gram stain reveals Gram (+) cocci. Consequent sputum and blood culture: Streptococcus pneumoniae. How do you treat him? Describe the factors in choosing appropriate antibiotic agents.

CEPHALOSPORINS

CEPHALOSPORINSCephalosporium acremonium

Cephalosporins / introductiony fungus Cephalosporium acremonium y source of the first members of this class y Advantageous over penicillins y A lot of combinations of modifications and alterations: large

availability of commercial cephalosporins.

structurey nucleus with two side chains y nucleus is 7-aminocephalosporanic acid y similar to the nucleus of penicillin y

-lactam ring is fused to a six-member dihydrothiazine ring instead of a fivemember thiazolidine ring

Advantages over penicillinsy RESISTANCE PRONE: intrinsically more resistant to

cleavage by -lactamases y MORE AVAILABILITY: two sites, R1 and R2, at which it can be modified:y large number of cephalosporins commercially available today.

y R1: ANTIBACTERIAL ACTIVITY y R2: PHARMACOKINETICS y modifications of the R1 side chain affect antibacterial activity

and alterations of the R2 side chain are associated with changes in the pharmacokinetic properties of these agents

Mechanism of effecty Attach to and inhibit penicillin-binding proteins (PBPs), thereby prevent

the appropriate synthesis of peptidoglycan. y peptidoglycan is a constituent of most bacteria y not active against certain species and strains of bacteria. y Why?y REMEMBER THE SIX PS:

1. 2. 3. 4. 5. 6.

PENETRATION PORINS PUMPS - EFFLUX PUMPS PENICILLINASES (really -lactamases, but that does not start with P!) PBPs PEPTIDOGLYCAN IS ABSENT

RESISTANCE TO CEPHALOSPORINS1.

Penetrationcephalosporins, like most -lactams, penetrate poorly into the intracellular compartment of human cells, so bacteria that for the most part reside in this compartment, such as Rickettsia and Legionella, are protected from them.

2/6 Porinsy Porinssome Gram-negative bacteria, such as Pseudomonas

aeruginosa, have porins in their outer membranes that do not allow passage of many cephalosporins into the periplasmic space.

3/6 Pumpsy Pumpssome bacteria, such as P. aeruginosa, use efflux

pumps to remove antibiotics from the periplasmic space.

4/6 Penicillinasesy Penicillinases (actually -lactamases)many Gram-negative

bacteria, such as Enterobacter and Citrobacter spp., make lactamases that degrade many cephalosporins.

5/6 PBPsy PBPssome bacteria, such as the enterococci and Listeria

monocytogenes, produce PBPs that do not bind cephalosporins with a high affinity.

6/6 Peptidoglycany Peptidoglycansome bacteria, like Mycoplasma and

Chlamydia, do not make peptidoglycan and therefore are not affected by the cephalosporins.

Generalizations of activity of cephalosporinsEach successive generation of agents has broader activity against aerobic Gram-negative bacteria. 2. With several important exceptions, these agents have limited activity against anaerobes. 3. The activities of these agents against aerobic Gram-positive bacteria are variable, but in general the first-generation agents have the strongest activity against these bacteria.1.

generationsGenerationFirst-generation Second-generation

Parenteral AgentsCefazolin Cefotetan,* cefoxitin,* cefuroxime

Oral AgentsCefadroxil, cephalexin Cefaclor, cefprozil, cefuroxime axetil, loracarbef Cefdinir, cefditoren, cefpodoxime proxetil, ceftibuten, cefixime

Third-generation

Cefotaxime, ceftazidime, ceftizoxime, ceftriaxone

Fourth-generation

Cefepime

FIRST GENERATION CEPHALOSPORINSCEFAZOLIN CEFADROXIL CEPHALEXIN

Structure of cefazolin

Strengthsy Activity against aerobic Gram-positive cocci such as

staphylococci and streptococci. y The R1 side chains of these agents protect their -lactam rings from cleavage by the staphylococcal -lactamase. y As a result, they are useful in the treatment of infections caused by many strains of Staphylococcus aureus.

Structure of cefazolin

Shortcomings / All cephalosporinsy ALL CEPHALOSPORINS cannot bind the PBPs of

methicillin-resistant S. aureus and methicillin-resistant Staphylococcus epidermidis or many highly penicillin-resistant Streptococcus pneumoniae (PRSP). y ALL CEPHALOSPORINS ARE INEFFECTIVE AGAINST MRSA AND MRSE OR PRSP PRSP. y ALL CEPHALOSPORINS also lack activity against Listeria monocytogenes and the enterococci.

Activity against Gram (-), Anaerobes and othersy Limited activity against aerobic and facultative Gram-

negative bacteria. y No protection from the -lactamases of most Gram-negative bacteria. y Some strains of Escherichia coli, Klebsiella pneumoniae, and Proteus mirabilis are susceptible. y Have moderate to poor activity against anaerobes, intracellular bacteria, and spirochetes.

Antimicrobial Activity of First Generation CephalosporinsBACTERIA Gram-positive bacteria BACTERIATYPE Streptococcus pyogenes Some viridans streptococci Some Staphylococcus aureus Some Streptococcus pneumoniae Some Escherichia coli Some Klebsiella pneumoniae Some Proteus mirabilis

Gram-negative bacteria

SECOND GENERATION CEPHALOSPORINSTrue cephalosporins. Ex: cefuroxime

The cephamycins: cefotetan and cefoxitin

cefoxitin

cefotetan

Common activity of true second generation ceph. And cephamycinesy Second generation cephalosporins are more active against:

aerobic and facultative Gram-negative bacteria, and: y more potent against E. coli, K. pneumoniae, and P. mirabilis than first-generation agents. y also active against Neisseria spp., and; y -lactamase-producing strains of Haemophilus influenzae (true cephalosporins: cefuroxime).

CEPHAMYCINSy Parent compound originally isolated

from the bacterium Streptomyces lactamdurans instead of the fungus Cephalosporium acremonium. y methoxy group in place of the hydrogen on the -lactam ring of the cephalosporin core.

Streptomyces sp.

Why include cephamycines with cephalosporins?y Pharmacologically and chemically similar agents with second

generation cephalosporin,i.e.; cefuroxime.

cefuroxime

cefotetan

cefoxitin

True cephalosporins and cephamycinsy Advantage of cefuroxime over cephamycines: y True cephalosporins are as active against the Gram (+) as

first generation cephalosporins. y Cephamycines (cefotetan and cefoxitin) have LIMITED activity against Gram (+)!

Additional methoxyy Additional methoxy: y Additional methoxy:

advantage over cefuroxime (at a cost!): enhanced stability to the lactamases of some anaerobes, such as Bacteroides fragilis.

diminished activity against staphylococci and streptococci because of decreased affinity for the PBPs of these bacteria.

Antimicrobial Activity of Second Generation CephalosporinsBACTERIA Gram-positive bacteria BACTERIATYPE Streptococcus pyogenes Some viridans streptococci Some Staphylococcus aureus Some Streptococcus pneumoniae

True cephalosporins have activity equivalent to first-generation agents Cefoxitin and cefotetan have little activity. Gram-negative bacteria Escherichia coli Klebsiella pneumoniae Proteus mirabilis Haemophilus influenzae Neisseria spp. Cefoxitin and cefotetan have moderate activity.

Anaerobic bacteria

THIRD GENERATION CEPHALOSPORINSCEFTRIAXONE, CEFOTAXIME, CEFTIZOXIME, CEFTAZIDIME

3rd Geny have moderate activity against aerobic Gram-positive

bacteria. y With the exception of ceftizoxime, they inhibit most strains of penicillin-susceptible S. Pneumoniae.y ceftizoxime is also unique in its ability to inhibit a significant

number of anaerobic bacteria, including many strains of B. fragilis.y Third-generation cephalosporins are also active against the

spirochete Borrelia burgdorferi.

aminothiazolyl group at R1ncreased activityy increased penetration

cefotaxime

Against;y E. coli, Klebsiella spp., Proteus

through the bacterial outer membrane, y increased affinity for PBPs, y increased stability in the presence of some of the plasmid-encoded lactamases of aerobic and facultative Gram-negative bacteria.

spp., Neisseria spp., and H. influenzae relative to the secondgeneration cephalosporins. y Many other strains of the Enterobacteriaceae, including Enterobacter spp., Citrobacter freundii, Providencia spp., Morganella morganii, and Serratia spp., also initially show susceptibility to thirdgeneration cephalosporins.

CAUTION! AmpC -lactamases!y Enterobacteriaceae harbor chromosomally encoded inducible

AmpC -lactamases that may allow the emergence of resistance during treatment. y Therefore; either do not use 3rd gen or use it with an additional beta-lactamase only if proven susceptible in vitro.

Activity against P. aeruginosay carboxypropyl group y dramatically increases y cefoperazone, also has

antipseudomonal activity y decreased affinity for the PBPs of staphylococci. y ceftazidime has enhanced activity against P. aeruginosa but limited activity against S. aureus

appreciable activity against P. aeruginosa, although it is not as active as ceftazidime.

cefoperazone

ceftazidime

ceftriaxoney Long half life: once a day!

Antimicrobial Activity of 3rd Generation CephalosporinsBACTERIA Gram-positive bacteria BACTERIATYPE Streptococcus pyogenes Viridans streptococci Many Streptococcus pneumoniae Modest activity against Staphylococcus aureus Escherichia coli Klebsiella pneumoniae Proteus spp. Haemophilus influenzae Neisseria spp. Some Enterobacteriaceae Borrelia burgdorferi

Gram-negative bacteria

Spirochetes

FOURTH GENERATION CEPHALOSPORINS

CHALLENGESy suffer from susceptibility to the chromosomally encoded

inducible AmpC -lactamases of many of the Enterobacteriaceae. y activity against P. aeruginosa is gained only at the expense of diminished antistaphylococcal activity.

CEFEPIMEy R1: AMINOTHIAZOLYL y R2: PYRROLIDINE (polar) y more rapid penetration through the outer membrane of

many Gram-negative bacteria, including P. aeruginosa. y binds at a high affinity to many of the PBPs of these bacteria. y relatively resistant to hydrolysis by Gram-negative lactamases, including the chromosomally encoded inducible AmpC -lactamases of the Enterobacteriaceae (?? Clinical controversy!) y Very limited anaerobic activity

Antimicrobial Activity of 4th Generation CephalosporinsBACTERIA Gram-positive bacteria BACTERIATYPE Streptococcus pyogenes Viridans streptococci Many Streptococcus pneumoniae Modest activity against Staphylococcus aureus Escherichia coli Klebsiella pneumoniae Proteus spp. Haemophilus influenzae Neisseria spp. Many other Enterobacteriaceae Pseudomonas aeruginosa

Gram-negative bacteria

TOXICITY OF CEPHALOSPORINSRash Urticaria anaphylaxis

Toxicity of cephalosporinsRARE SIDE EFFECTS y These are relatively SAFE pharmaceuticals. y RARELY: immediate hypersensitivity:y Rash y Urticaria y Anaphylaxis

OTHER MORE RARE SEy reversible neutropenia, y thrombocytosis, y hemolysis, y diarrhea, and y elevated liver function tests.

y Those who developed the

above due to penicillins are recommended not to use cephalosporins.

Special toxicityCefotetan & cefoperazony Hypoprothrombinemia* y +alcohol => disulfiram5-10 times more acetaldehyde in blood: Hangover symptoms emphasized!

like effects due to methylthiotetrazole moiety at R2 of these agents

methylthiotetrazole

Brief note: hypoprothrombinemiay Deficiency

of prothrombin (Factor II) results in impaired blood clotting, y increased physiological risk for bleeding, y especially in the gastrointestinal system, cranial vault, and superficial integumentary system.y Integumentery system: the

organ system that protects the body from damage, comprising the skin and its appendages (including hair, scales, feathers, and nails).

Brief note: Prothrombin and thrombiny Prothrombin (factor II of the coagulation

cascade) is a critical protein in hemostasis. y Activated factor Xa converts prothrombin to thrombin. y Thrombin is a potent protease. Its most important function is the cleavage of fibrinogen to create insoluble fibrin. Crosslinking fibrin monomers stabilize the fibrin clot. Factor XIIIa, activated by thrombin, carries out this function. y Decreased levels of prothrombin (hypoprothrombinemia) leads to mucocutaneous bleeding and hemorrhage due to lack of prothrombin.

Brief note: Coagulation cascade

Ceftriaxone & biliary sludgey Excreted by biliary excretion: y High doses: biliary sludge. y Sludge: residual, semi-solid material left from

industrial wastewater, or sewage treatment processes. y Biliary sludge: A mixture of microscopic particulate matter in bile that occurs when particles of material precipitate from bile. y Biliary sludge, however, may cause intermittent symptoms and, on occasion the particles may grow in size and become larger gallstones: nausea and vomiting, and Pancreatitis

SUMMARY - CEPHALOSPORINSy Vary in activity. y First-generation agents have STRONG activity against

aerobic Gram-positive bacteria. y Second-generation agents have modest activity against aerobic Gram-positive, aerobic Gram-negative, and (in some cases) anaerobic bacteria. y Third-generation agents have STRONG activity against aerobic Gram-negative bacteria. y Fourth-generation agents have ESPECIALLY ENHANCED activity against aerobic Gram-negative bacteria.

Lets reviewy 1st Gen

Gram (+) and STRONG! y 2nd Gen Gram (+), Gram (-), and some anaerobic but MODEST. y 3rd Gen Gram (-) and STRONG! y 4th Gen Gram (-) and ESPECIALLY ENHANCED (ENHANCED!

MNEMONIC : Pain in the neck!GENERATION MNEMONIC First PECK Second Third Fourth PIN NECK PEN IN NECK PAEN IN NECK COVERAGEProteus mirabilis Escherichia coli Klebsiella pneumoniae + Haemophilus influenzae +Neisseria spp. + some Enterobacteriaceae + Pseudomonas aeruginosa + many Enterobacteriaceae

WHICH CEPHALOSPORIN TO USE WITH WHICH MICROORGANISM? RATIONAL DRUG USE.

CARBAPENEMSTHE LAST LINE OF DEFENSE: these guys mean bad business! IMIPENEM(/CILASTATIN), MEROPENEM & ERTAPENEM IMIPENEM(/ ),

Carbapenemsy mipenem + silastatin sodyumy Tienam (flakon i.v. ve i.m.)

y Meropenemy Meronem (flakon i.v.)

y Ertapenemy Invanz (flakon i.v. ve i.m.)

y Doripenem y AZTREONAM

Differences of carbapenems from penicillins: the structureThe sulfur is replaced by a methylene group. 2. The ring contains a double bond.1.Penicillin structure

Carbapenem structure

Extra activities of carbapenems- WHY?y INCREDIBLE BROAD SPECTRUM OF ACTIVITY!

1.1.

VERY SMALL MOLECULESThese molecules are quite small and have charge characteristics that allow them to utilize special porins in the outer membrane of Gram-negative bacteria to gain access to the penicillin-binding proteins (PBPs).

2.1.

BETA-LACTAMASE RESISTANCETheir molecular structure makes them resistant against lactamases

3.

AFFINITY TO PBPs of BROAD RANGE OF BACTERIA

SO WHAT?y Because of the aforementioned

three properties, carbapenems are; Adept at gaining access to periplasm! 2. resisting destruction by lactamases that reside there, and 3. binding to PBPs to cause bacterial cell death.1.

Resistance

Resistancey Pseudomonas aeruginosay Mutation of porins porins: y Carbapenems cant go through mutated porins. y Increasing the number of efflux pumps: pumps y Carbapenems cant accumulate in the periplasm.

y Enterococcus faecium and methicillin-resistant

staphylococci:y produce altered PBPs that do not bind these carbapenems

y Some bactaria produce extremely powerful -lactamases that

are capable of cleaving carbapenems.

IMIPENEMy Structural simplicity: y no R1! y Derived from y StreptomycesBeta lactams

cattleyaCarbapenem structure

imipenem

dehydropeptidase I & CILASTATINy Rapidly destroys imipenem in the kidney because of lack of

R1 in its structure. y Therein comes CILASTATINcilastatin

imipenem

CILASTATIN

IMIPENEM + CILASTATINy Administered together to inhibit dehydropeptidase I from

metabolizing imipenem. y Active against many species of pathogenic bacteria! y Most Streptococci: including Penicillin resistant Streptococcus pneumoniae. pneumoniae y Most Staphylococcusy But not against MRSA!

y Many Gram negativey P. aeruginosa

y the highly resistant Enterobacteriaceaey Enterobacter Citrobacter Enterobacter,

y excellent anaerobic coverage y But not against Clostridium difficile!

Activity summaryy Active against most bacteria resistant to

penicillins and cephalosporins. y Wide antibacterial spectrum y Surpasses activity of 3rd Gen Cephalosporins. y Not active against MRSA! y JUST LIKE ALL OTHER BETA LACTAMS!

mipenem + Silastatin SodyumEndikasyon (i.v.) (LKEM ZDE)Alt solunum yolu enf. y Staphylococcus aureus, y Escherichia coli, y Klebsiella sp., y Enterobacter sp., y Haemophilus influenzae, y Haemophilus parainfluenzae, y Acinetobacter sp., y Serratia marcescens. drar yolu enf.y Enterococcus faecalis, y S. aureus, y E. coli, y Klebsiella sp., y Enterobacter sp., y Proteus vulgaris, y Providencia rettgeri, y Morganella morganii y P. aeruginosa.

mipenem + Silastatin SodyumEndikasyon (i.v.) (LKEM ZDE)Kar n ii enf. y Enterococcus faecalis, y S. aureus, y Staphylococcus epidermidis, y E. coli, y Klebsiella sp., y Enterobacter sp., y Proteus sp., y Morganella morganii, y P. aeruginosay Citrobacter sp., y Clostridium sp., y Bacteroides sp. (B. fragilis dahil) y Fusobacterium sp., y Peptococcus sp., y Peptostreptococcus sp., y Eubacterium sp., y Propionibacterium sp., y Bifidobacterium sp.

mipenem + Silastatin SodyumEndikasyon (i.v.) (LKEM ZDE)Kad n hastal klar ile ilgiliy Enterococcus faecalis; y S. aureus, y S. epidermidis, y Streptococcus agalactiae y Enterobacter sp., y Bifidobacterium sp., y Bacteroides sp. (B. fragilis y y y y

(group B streptococcus), y E. coli , y Klebsiella sp., y Proteus sp.

dahil) Gardnerella vaginalis; Peptococcus sp., Peptostreptococcus sp., Propionibacterium sp.

MEROPENEMy Resistant to dehydropeptidase I: R1 has a methyl group. y spectrum of activity is essentially the same as imipenem.

ERTAPENEMy not cleaved by renal dehydropeptidase because of methyl in

R1 (just like imipenem). y R2 side chain: is less active against aerobic Gram-positive bacteria, P. aeruginosa, and Acinetobacter spp. than the other carbapenems! y Once a day.

CARBAPENEMS AND ADVERSE EVENTS.y Nausea, vomiting, diarrhea, rash, and

drug fever y Seizures y Contra-ind: pre-existing central nervous system disease and with renal insufficiency

Antimicrobial Activity of CARBAPENEMSBACTERIA Gram-positive bacteria BACTERIATYPE Streptococcus pyogenes Viridans group streptococci Streptococcus pneumoniae Modest activity against Staphylococcus aureus Some enterococci Listeria monocytogenes Haemophilus influenzae Neisseria spp. Enterobacteriaceae Pseudomonas aeruginosa Bacteroides fragilis Most other anaerobes

Gram-negative bacteria

Anaerobic bacteria

Faecium or faecalis?y Strains of Enterococcus faecalis that are

susceptible to penicillin are also susceptible to carbapenems y (except ertapenem). y Enterococcus faecium, however, is resistant to all carbapenems!

MONOBACTAMSThe lonely cowboy: aztreonam. DOES ONLY ONE THING BUT DOES IT VERY WELL!

AZTREONAMy kills aerobic Gram-

AZACTAM (art k ithal edilmiyor)

negative bacteria. y the aminothiazolyl group that so dramatically improves the aerobic Gram-negative coverage of third-generation cephalosporins.

CEFOTAXIME

AZTREONAM

Aminothiazolyl group & aztreonamy Binds quite well to the penicillin-binding proteins (PBPs) of

aerobic Gram-negative bacteria. y stable against many of the -lactamases. y excellent activity against Neisseria and Haemophilus spp. y intermediate activity against Pseudomonas aeruginosa

Pseudomonas aeruginosa

Haemophilus influenzae as seen in a Gram (-) technique

Neisseria meningitidis

Some shortcomingsy does not bind the PBPs of Gram-positive or anaerobic

bacteria.y Can not cure infections by Gram (+) or anaerobic bacteria.

y Resistance: the Enterobacteriaceae and P. aeruginosay changes in the permeability of the outer membrane y destruction by -lactamases

Pseudomonas aeruginosa

Toxicityy Aztreonam is very safe: y NOT associated with nephrotoxicity. y a renal-sparing alternative to the aminoglycosides (both

against Gram negative bacteria). y Penicillin allergies? y Safe to use aztreonam in patients with penicillin allergies.

Antimicrobial Activity of Aztreonam

BACTERIA Gram-negative bacteria

BACTERIATYPE Haemophilus influenzae Neisseria spp. Most Enterobacteriaceae Many Pseudomonas aeruginosa

VANCOMYCIN

Historyy discovered when a missionary from Borneo y y y y

sent a soil sample to his friend, who was an organic chemist at Eli Lilly and Company. The soil sample turned out to harbor a bacterium that made a compound with potent activity against Gram-positive bacteria. Eventually the compound was purified and named vancomycin, which is derived from the word vanquish.Ref: Griffith RS. Vancomycin usean historical review. J Antimicrob Chemother 1984;14(Suppl D):15.

Structure of vancomyciny A glycopeptide! Not a beta-lactam. a peptide with sugar

moieties attached to it. y Intact peptides are poorly absorbed in the GI tract.

Its structure helps it to bind!y Similarity to beta lactams: preventing synthesis of the cell

wall. y binds to the D-alanyl-D-alanine portion of the peptide side chain of precursor peptidoglycan subunits.

Mechanism of penicillin-binding protein (PBP) inhibition by -lactam antibiotics.A. PBPs recognize and catalyze the peptide bond between two alanine subunits of the peptidoglycan peptide side chain. B.The -lactam ring mimics this peptide bond. Thus, the PBPs attempt to catalyze the -lactam ring, resulting in inactivation of the PBPs.

Bulky molecule!y Vancomycin binds to the D-alanyl-D-alanine dipeptide on the

peptide side chain of newly synthesized peptidoglycan subunits, preventing them from being incorporated into the cell wall by penicillin-binding proteins (PBPs).

A glycopeptidey poorly absorbed in the gastrointestinal

tract: only i.v forms available. y Clostridium difficile: effective against diarrhea by C. difficile.y ONLY ORAL FORMS not absorbed thru

GI track, thats why!y Not able to pass through PORINS: not

effective against Gram (-) y But very effective against Gram (+).y Question: Can you name an antibiotic

which is the opposite?Clostridium difficile associated collitis

Active against:y nearly all staphylococci and streptococci,

including methicillin-resistant staphylococci and strains of penicillinresistant Streptococcus pneumoniae. y Listeria monocytogenes: susceptible in vitro but not so much in vivo! CAUTION! y good activity against anaerobic Grampositive bacteria, including C. difficile.

Streptococcus pneumoniae

Listeria monocytogenes

C. difficile

Antimicrobial Activity of VancomycinBACTERIA Gram-positive bacteria BACTERIATYPE Staphylococcus aureus (CAUTION!) Staphylococcus epidermidis Streptococcus pyogenes Viridans group streptococci Streptococcus pneumoniae Some enterococci (exceptVRE) Clostridium spp. Other Gram-positive anaerobes

Anaerobic bacteria

RESISTANCE: VREy Vancomycin Resistant Enterococci (VRE): the D-alanyl-D-

alanine dipeptide is replaced with D-alanyl-D-lactate, which is not recognized by vancomycin. Thus, the peptidoglycan subunit is appropriately incorporated into the cell wall.

Gene transfer S. aureusy Unfortunately, the gene clusters that encode this activity in

enterococci are transferable and have already been found in Staphylococcus aureus.Thus, it is anticipated that vancomycin resistance will occur with increasing frequency in staphylococci as well.

Toxicityy Hearing loss esp. w/

aminoglycosides y Red man syndromey develop pruritus and an

erythematous rash on the face, neck, and upper torso.y Neutropenia: dangerously or

abnormally low count of neutrophils in the blood stream.Atypical white blood cell count per cubic mm of blood should be be neutrophils 3650, monocytes 430, lymphocytes 2500, eosinophils 150 and basophils 30. Neutrophils kill and absorb invasive microorganisms. Neutropenia occurs when the neutrophils count measures less than 2000 cells per cubic mm.

DAPTOMYCINa novel cyclic lipopeptide antibiotic

Daptomycine the first of a new classy First of a new class: THE CYCLIC LIPOPEPTIDES y Rapidly bactericidal-concentration dependent. y Active against Gram(+) pathogens. y Unique mode of action.y No cross-resistance with other antimicrobial classes!

y 90% protein (92% albumin) bound.y Once a daily IV

y Poor efficacy in pulmonary infections. y RESISTANCE reported! (during clinical trials

PHASE II)y E faecalis, S aureus, and among clinical isolates of

MRSA

Chemical structurey The lipophilic tail: lipid portion of this

drug inserts into the bacterial cytoplasmic membrane, where it forms an ion-conducting channel. y Ions escape from the bacterium. y Bacterium dies.

Daptomycin is a 13-member amino acid cyclic lipopeptide compound that contains a watersoluble hydrophilic core with a lipophilic tail. The lipophilic tail exerts itself into the cytoplasmic membrane of the gram-positive cell wall.

Mechanism of actiony The acyl tail inserts itself into the cytoplasmic membrane. y forms a channel that causes depolarization of the membrane. y efflux of potassium (and possibly other) cytoplasmic ions, thus

inhibiting macromolecular synthesis and leading to cell death.

Peculiarity: Stealth!y Concentration dependent. y Rapid. y Unlike that of many other antibiotics. y Depolarization of acyl tail is correlated with daptomycin's

bactericidal activity. y Why stealth? Leaves a ghost cell behind completely drained. y But daptomycin DOES NOT rupture the cell wall!

ActivityActive against: y many aerobic Gram-positive bacteria, y including highly resistant strains such as y methicillin-resistant Staphylococcus aureus, y penicillin-resistant Streptococcus pneumoniae, and y some vancomycin-resistant enterococci! y GREAT! Unfortunately not active against:y Gram-negative organismsy Cannot reach the

cytoplasmic membrane through the Gram(-) outer membrane.y Pneumonia: poor activity

in lungs!y Pulmoner srfaktan

taraf ndan etkisizle tirilir.

Ek note: Pulmoner srfaktan nedir?y Akci er s v lar n n yzey gerilimlerini azaltarak solunum

yollar n n a k kalmas n sa lamaya yard mc olan do al maddelere benzeyen yzey aktif bile iklerdir. y Eksojen pulmoner surfaktanlar, prematre bebeklerde neonatal solunum s k nt s sendromunda 100 200 mg fosfolipit / kg (do um a rl ) olarak, ilk doz te histen sonra en k sa zamanda uygulan r. Entbe bebeklerde endotrakeal tp vas tas yla mekanik solunuma solsyon olarak verilebilir. y Ba l calar : beraktant, kolfoseril palmitat, poraktan alfa, s r akci er srfaktan fosfolipid fraksiyonu, kalfaktan, lusinaktan, pumaktant, sinapultit

Pulmoner srfaktanlar neden daptomycini tutarlar?y Bunlar n yap s nda iki temel yap vard r: y Fosfatidilgliserol y Dipalmitoilfosfatidilkolin y Bu iki madde Gram (+) hcre membran nda da vard r! y Daptomycin bu maddelerin iine girer, hapsolur ve etkisini

gsteremez.

Antimicrobial Activity of DaptomycinBACTERIA Gram-positive bacteria BACTERIATYPE Streptococcus pyogenes Viridans group streptococci Streptococcus pneumoniae Staphylococci Enterococci

Anaerobic bacteria

Some Clostridium spp. Clostridium difficile, Clostridium perfringens,

General IndicationPrimarily used for: y SKIN y SOFT TISSUE infections. approved in 2003 in the U.S. for the treatment of complicated SSSIs caused by S. aureus, including MRSA, as well as Streptococcus pyogenes, Streptococcus agalactiae, Streptococcus dysgalactiae subsp. equisimilis, and Enterococcus faecalis (vancomycinsusceptible isolates only). Recently, the label for daptomycin was extended to include the treatment of bacteremia, including rightsided endocarditis caused by S. aureus.** Cubist Pharmaceuticals. Cubicin (daptomycin for injection) prescribing information. 2006. Lexington, MA.

Why do we need daptomycin?y MRSA: First choice of treatment is

VANCOMYCIN, buty Needs monitoring of dosages & y Can cause nephrotoxicity!

y RESISTANCE!y vancomycin-resistant S. aureus (VRSA) y vancomycin-resistant enterococci (VRE) y vancomycin-intermediate S. aureus (VISA)

y Commonly used linezolids disadvantage:y one may need to limit the duration of therapy due to

hematologic adverse events (suppression of blood cell counts).

Dis- and Advantages of daptomycinAdv y No other agent detected or known (yet?!) that is resistant.y There is very little data of

Disadvy Effectiveness is dependent

on free biologic calcium.y To let the efflux of K+

losing susceptibility.*

y No other agent detected or

y Effectiveness can be

known that cause transresistance (like theVRE from VRS aureus). y Once a daily dose y Less renal deficiency causing**

hindered by albumin concentrations. y Dose dependence and combinations still under investigation.

Toxicityy Relatively safe and well tolerated. y High doses y Phlebitis y Rash y GI intolerance

reversible myopathy.