Bacterial Pathogenesis

Kunle Kassim, PhD, MPH Professor, Microbiology August, 2010

URGENT!!!!
It is important for you to review the lecture powerpoint on Host/Bacteria Interactions from first yearbefore coming to class for this lecture.

Objectives
Review the diversity and significance of bacterial flora in the maintenance of immunity Review the various types of flora distruption and health consequences to the host. Review determinants of pathogenicity Discuss the roles of endotoxin and exotoxins in specific instances of bacterial pathogenesis Illustrate invasiveness and dissemination of bacterial pathogenesis with Salmonella and Shigella pathogenesis. Discuss the different ways that bacterial pathogens exert damage and injury to the host, using cystic fibrosis, lyme disease and bacterial urethritis as illustrations Present the different types of host defenses, including the constitutive elements of innate and adaptive immunity, humoral and cellular immunity, inflammatory and acute phase responses. Describe the emerging patterns and serious significance of nosocomial infections Discuss the nature, usefulness and schedule of bacterial vaccines, particularly in preventing childhood infectious diseases.

Normal Bacterial Flora Bacterial Virulence Factors Mechanisms of Pathogenicity

Host Defense Mechanisms Selected Bacterial Diseases

(Review lecture materials on Bacteria/Host Intreactions)

Distruption of Normal Flora

Trauma - appendix rupture, dental extraction, auto accidents, gun violence Flora displacement/contamination - UTI, bacterial vaginosis Excessive antibiotic use - vaginal candidaisis, pseudomembranous colitis by Clostridium difficile

Distruption of Normal Flora

UTI
Tumors Bladder incontinence Ureteric reflux Poor hygiene

E. coli infections

Determinants of Pathogenicity
Ports of entry Modes of transmission Bacterial adherence Invasiveness/Dissemination Pathological damage Endotoxin / Exotoxins Fever / Disease onset Host defenses Evasion of host defenses Siderophore production Plasmids Bacterial vaccines (Review last years lecture materials on Bacteria/Host Interactions)

Toxins
Endotoxin (Lipid A component of LPS)
-Unlike exotoxins, lipid A is not antigenic and cannot be converted to a toxoid; responsible for gram negative septicemia, with a fatality rate of 25-50 percent in USA

Exotoxins
-most are polypeptides, are antigenic and can be converted to toxoids

Endotoxin and Septicemia

Fever

Activation of coagulation process Depression of RES Vascular collapse

Lipid A, Coagulation and RES

Lipid A activates clotting mechanism, with formation of fibrin Fibrin may clog small blood vessels, causing intravascular coagulation, followed by shock Lipid A may inhibit macrophages from degrading fibrin polymers trapped in blood vessels

Lipid A and Vascular Collapse

Lipid A activates macrophages to release TNF-, which causes increased vascular permeability and dilatation This causes low blood pressure (hypotension), impairs blood flow to vital organs (kidneys, liver, lung, brain), followed by shock, multiple organ failure and death

Neisseria species
Are all gram negative cocci Oxidase, catalase positive Multiply intracellularly Neisseria meningitidis Neisseria gonorrhea Neisseria sicca

Neisseria meningitidis
Gram negative diplococci Infection from aerosol transmission in close contacts Polysacharide capsule is antiphagocytic LPS, capsule and sIgA protease are major virulence factors Meningitis, fever, pneumonia, meningococcemia with hemorrhagic lesions are major clinical manifestations Prevented by immunization with polysacharide-protein conjugate vaccine

Meningococcemia

Death from Neisseria septicemia and meningitis

Neisseria urethritis
Gonorrhea with purulent discharge Disseminated infections via blood to skin, joints Ophthalmia neonatorium (acquired eye infection)

Complications of Gonococcal Infection

Skin lesion

Septic arthritis

Ophthalmia neonatorium

Neisseria gonococcus / Chlamydia Urethritis Differentiation

Bacterial Classification and Pathogenesis

Meningitis
(Strep pneumo, H. influenzae, N. meningitidis)

Exotoxins
Enzymatic lysis alpha toxinClostridium perfringes Pore formation alpha toxinStaph aureus Protein synthesis inhibition diphtheria & shigella toxins Nerve-muscle transmission-inhibition tetanus toxin-spastic paralysis botulinum toxin-flaccid paralysis

Modes of Action of Selected Exotoxins

Cholera (Vibrio cholerae)

Clostridium and Bacillus

Identification of species based on spore location and oxygen metabolism: - C. tetanus - C. botulinum - C. perfringes - C. difficile - B. anthracis - B. cereus

Spore Location for Species ID

Tetanus
(Clostridium tetani and skeletal muscle flexion)

Bacillus anthracis (potential bioterrorism agent, 2001)

Gram Stain of B. anthracis in Lung Exudate of an Anthrax Patient

Bacterial Classification and Pathogenesis

Staphylococcus and Streptococcus

Tetrad cocci of Staph aureus (MRSA) String-like cocci of Strep pneomoniae/ Strep viridans Enterococcus faecalis MDR-Strep/Entero

Staphylococcus aureus
Diseases Toxin-mediated food poisoning, toxic shock syndrome; cutaneous impetigo, folliculitis, wound infections, pneumonia, osteomyelitis, septic arthritis Virulence Factors Capsule, protein A, cytotoxins/hemolysins, enterotoxins, exfoliative toxin; coagulase/catalase/beta-lactamase enzymes Methycillin-resistant Staph aureus (MRSA) (antibiotic resistant strain, serious problem in hospitals, in the US military, among athletes)

Scalded skin syndrome due to Staphylococcus aureus exotoxin

Gram Stain of Strep viridans in exudate of cardiac valves

Bacterial endocarditis from a dental extraction

E. coli Pathogenesis
Diseases Gastroenteritis, bacteremia, UTI, cystitis, pyelonephritis, neonatal meningitis, intraabdominal infections Virulence Factors Capsule, LPS, shiga toxins, hemolysins, siderophores (enterobactin, aerobactin), R-plasmid

Invasiveness / Dissemination
Exotoxins and Extracellular enzymes Extracellular enzymes
_ Hyaluronidase dissolves connective tissue Collagenase hydrolyses muscle connective tissue Streptokinase lyses blood clots Phospholipases damage cell membranes Lecithinase damages cell membranes Staphylokinase (fibrinolysin) dissolves fibrin clots Hemolysins lyse erythrocytes and white blood cells

Shigella/Salmonella Invasive & Dissemination Mechanisms

Shigella Invades M cells w 4 invasion proteins (IpaA,B,C,D) Replicates in host cell cytoplasm Induces apoptosis of phagocytes Produces cytotoxin (60s ribosome protein inhibitor) Invades deeper tissues, but not blood circulation Causes diarrhea, dysentery Low infective dose (100 cells) Salmonella Invades w invasion proteins Replicates in acidic host cell vacuole Replicates and transported by macrophages in blood circulation Invades other systemic organs, but not deeper GI tissues Causes diarrhea, enteric fever High infective dose (>I million cells)

Salmonella/Shigella Invasive & Mechanisms

Cystic Fibrosis
Genetically inherited disorder occurring in 1 of 2500 live Caucasian births Caused by pancreatic insufficiency, abnormal sweat electrolyte concentrations, viscid bronchial secretions Bronchial secretions lead to stasis in lungs and disposition to infections ( Staph aureus, H. influenzae, Pseudomonas aeruginosa most virulent and antibiotic resistant) and pneumonia

Cystic Fibrosis

Lyme Disease

Lyme Disease
Caused by Borrelia burgdorferi Transmitted by hard ticks (Ixodes species) Human infections most prevalent in summer months in Europe and USA Accompanied by fever, headache, myalgia, lymphadenopathy, skin lesions (erythema chronicum migrans) Neurological complications (meningitis, encephalitis, peripheral neuropathy) Cardiological (heart block, myopericarditis) Arthralgia, arthritis (immune complex mediated) Treatable with penicillin or tetracycline at early stage

Lyme Disease (rash of erythema chronicum/ inflammation)

Lyme Disease

Host Defenses
Barriers to Infection Innate and adaptive immunity Humoral and cellular immunity Phagocytosis Complement activation Inflammation Acute phase response Hypersensitivity Reactions

Barriers to Infection

Innate and Adaptive Immunity

Antibody Responses to Infections

Various Roles of Macrophages

Macrophage and Neutrophil

Host immune responses to bacterial infections

Cytokine and Antibody Networks in Bacterial Infections

Inflammation
Consequence of a microbial infection Recruitment of inflammatory cells (neutrophils, macrophages, basophils, eosinophils) and endogenous mediators (complement, prostaglandin E2) to sites of infection Production of inflammatory cytokines ( IL-1, IL-6, TNF-) to induce acute-phase response May be accompanied by antigen neutralization and cytotoxicity

Stages of Inflammation
Tissue Damage Vasodilation Exudation Endothelial adherence Diapedes (phagocyte migration) Tissue repair

Inflammation from Lyme Disease (rash of erythema chronicum)

Induction of acute phase proteins in response to an infection

Acute Phase Response

Also a response to infection as with inflammation Triggered by IL-1, IL-6, TNF-, inflammation, prostaglandin E2, interferon Induce production of acute phase proteins (C-reactive protein, complement components, coagulation proteins) Reinforce innate defenses (complement activation, phagocytosis) against infection, but excessive production during sepsis by LPS can lead to shock What are other acute phase responses???

Hypersensitivity Reactions to Bacterial Infections

Antigenic Variation
Periodic changes of surface antigens by the organism allows it to bypass and not be affected by the host immune responses. This occurs in HIV infection, Lyme disease and trypanosomiasis.

Stages of a Disease

Hospital-acquired (Nosocomial) Infections

Pennsylvania 2004 Nosocomial Infections

Sites of Antibiotic Activities

Immunization
Passive maternal antibodies thr placenta and mothers milk Active -- natural exposure to microbes - exposure to vaccines Live vaccine attenuated Mycobacterium bovis Inactivated toxoid tetanus vaccine Inactivated killed typhoid vaccine Subunit capsular polysacharides (poor immunogens) -Hemophilus influenzae b, Neisseria meningitidis, Streptococcus pneumoniae Salmonella typhi Conjugate vaccines polysacharide units conjugated to protein molecules

Childhood Immunization Schedule

Case Studies

CASE 1: URINARY TRACT INFECTION Mr. Hamilton, a 69-year old man, underwent a transurethral prostatectomy for cancer of the prostate. Because of concern about postoperative bleeding during urination, the surgeons placed a Foley catheter into his bladder. Three days later, Mr. Hamilton developed a urinary tract infection with low-grade fever, some pain, and pyuria. Laboratory cultures yielded 3 x 105 colonies of Escherichia coli per ml of urine. The organisms were resistant to all tested antibiotics except for aminoglycosides. Within 2 days, Mr. Hamilton developed bacteremia with hypotension and shock. His doctor eventually controlled his bacteremia with gentamycin therapy.

Questions (#1)
I. What was the source of Mr. Hamilton's infection? 2. What bacterial component induced his fever, hypotension and shock and how? 3. List three characteristics of aminoglycosides and their modes of action. 4. What other organism could have been isolated from his infection site? Give reasons for your choice of organism.

CASE 2: NOSOCOMIAL WOUND INFECTION

Ms. Wilson, an 85-year-old woman with rheumatic heart disease, underwent a mitral valve replacement along with surgery for a coronary artery bypass graft. Her postoperative course was complicated by bleeding in the mediastinum, which required more surgery .She did well after these operations and was discharged after 12 days. Three weeks later, Ms. Wilson, noticed some purulent drainage along the wound site on her chest. She continued to have pain but did not tell her family, assuming that the pain was related to her healing process. When she returned to see the surgeon I month later, she reported her pain and low-grade fever. The surgeon noted that there was considerable drainage at the wound site. Probing the wound, he noticed a lot of pus. Ms. Wilson was hospitalized again for radical debridement (cleaning) of her chest wound. Cultures of the pus yielded Staphylococcus epidermidis. She was treated intravenously with vancomycin for 6 weeks and her wound was debrided, with the wires in her sternum removed. At the end of this period, she required a plastic surgical procedure and a muscle flap to close the wound. After 2 more months of hospitalization, she was discharged and continued her convalescence at home.

Questions (#2)
I. Describe the bacterial pathogenesis of rheumatic heart disease., including the organisms that may be associated with its causation. 2. What was the source of her postoperative wound infection? 3 .What is the mode of action of vancomycin and why was it the drug of choice for Mrs. Wilson's wound infection? 4. In the absence of vancomycin, what other antibiotic(s) would you choose to clinically manage the wound infection? Describe its mode of action.

Case # 4
Simo da Silva, 36-year-old male resident of New Jersey, developed joint and muscle pains and an expanding erythematous skin lesion on his left leg shortly after his summer vacation. A week later, he started experiencing severe headache, neck stiffness and photophobia. He also noticed multiple secondary annular skin lesions three weeks later. Blood analysis reveled a high titer of IgM antibodies against a Gram negative spirochete, but the IgG response was negative. Ten months after the resolution of the initial symptoms, Mr .da Silva had severe neuritic pain on the skin of his abdomen within the distribution of the T8 through T 11 dermatomes. This symptom was followed by intermittent joint pains, which occurred in one joint at a time for several days, followed by longer pain- free periods. During the second year of illness, the patient had a sudden onset of severe swelling of one knee and then the other. Synovial fluid analysis revealed numerous white cells, and his antibody response was high for IgG and low for IgM. His immunogenic profile showed that he had Ill-A-DR4 and Ill-A-DR+ specificities. The swelling of the knees remained for about one year, but then subsided.

Questions (#4)
I. How did Mr. da Silva acquire the spirochete infection and what is your identification of the organism? 2. What antibiotics would you use to treat his initial and late diseases? Is this a case of acute or chronic infection or disease? 3. What bacterial and immune factors are responsible for his intermittent joint pains and knee swelling? 4. Explain the initial high IgM and no IgG titers in the early stage of the disease and the low IgM and high IgG titers in Mr .da Silva late disease stage. 5. What type of white cells ( T cells, B cells, neutrophils, basophils, macrophages ) were found in the synovial fluid? 6. What does his immunologic profile ofHLA-DRA4 and HLA-DR2 specificities mean?

Case #5
Three young males were brought to Howard University Hospital Emergency Clinic over the course of two days. They were diagnosed with bacterial meningitis, but their cerebrospinal fluid (csf) cultures yielded three different organisms. They were all lethargic with fevers of 101 to 104o F, white blood cell (wbc) counts of over 15,000 cells/l, mostly neutrophils. One of the bacterial isolates was identified as Streptococcus pneumoniae, the second as Hemophilus influenzae tybe b and the third as Neisseria meninigitidis.

Questions (#5)
What bacterial culture chracteristics and laboratory methods were used to separately identify the three organisms? What is the pathogenic bacterial structure that is common to the three organisms and how is its composition different for each organism? Describe the pathway(s) that were taken by the organisms to get into the patients csf. What are the epidemiological characteristics of the meningitis caused by each organism? What antibiotics of choice would you use to treat the patients? Give reasons for your choice. What are the modes of action for your choice of antibiotics? What types of resistance mechanism may be used by each of the organisms to inhibit the activity of your selected antibiotics?

Home-Work Exercise
List organisms that may be associated with the following conditions 1. Bacteremia 2. Endocarditis 3. Meningitis 4. Pharyngitis 5. Pneumonia 6. Conjunctivitis 7. lntra-abdominal abscess 8. Gastroenteritis 9. Urinary Tract infections 10. lmpetigo 11. Cellulitis 12. Sepsis

Reading References
Chapters 9-13, 18 , 47 in Medical Microbiology, 6th edition by Patrick Murray et al, Mosby, Inc., 2009 Chapters 8 -10 in Medical Microbiology, 3rd edition by Cedric Mims, et al, Mosby, Inc., 2004. Chapters 6, 7, 8 and 9 in Mechanisms of Microbial Diseases, 3rd edition by Moselio Schaechter, et al, William & Wilkins, 1998.