Bacterial Infections Lecture Notes PDF

6/27/2024 CHAPTER 20 Serological and Molecular Detection of Bacterial Infections PREAMBLE PowerPoints are a general overview and are provided to help students take notes over the video le...

6/27/2024 CHAPTER 20 Serological and Molecular Detection of Bacterial Infections PREAMBLE PowerPoints are a general overview and are provided to help students take notes over the video lecture ONLY.  PowerPoints DO NOT cover the details needed for the Unit exam Each student is responsible for READING the TEXTBOOK for details to answer the UNIT OBJECTIVES Unit Objectives are your study guide (not this PowerPoint) Test questions cover the details of UNIT OBJECTIVES found only in your Textbook! 1  6/27/2024 CHAPTER OVERVIEW Host–microbe relationships Bacterial structure and virulence factors Immune defenses against bacteria and escape Laboratory detection of bacterial infections Clinical manifestations and laboratory detection of selected bacterial infections Group A streptococci Helicobacter pylori Mycoplasma pneumoniae Rickettsial infections HOST–MICROBE RELATIONSHIPS Host and microbes live together long term Symbiotic Indigenous microbiota Commensalistic No benefit or harm to either organism Mutualistic Both host and microbes benefit Parasitic Microbes cause harm to the host 2  6/27/2024 INFECTIVITY, PATHOGENICITY, AND VIRULENCE Infectivity Pathogenicity Virulence Organism’s Ability of Extent of ability to an pathology establish an organism to caused by infection cause an disease organism when it infects a host STRUCTURAL COMPONENTS OF BACTERIA 3  6/27/2024 BACTERIAL VIRULENCE FACTORS Endotoxin The lipid A portion of LPS in gram-negative cell walls Powerful stimulator of cytokine release Pili Adherence to host cells; resistance to phagocytosis Flagella Adherence to host cells; motility Capsule Blocks phagocytosis, antibody attachment, C’ Exotoxins Potent toxic proteins released from living bacteria Neurotoxins, cytotoxins, enterotoxins IMMUNE DEFENSES AGAINST BACTERIA Innate defenses Adaptive defenses Intact skin and mucosal Antibody production surfaces (barriers to Binding of C’, entry) opsonization, Antimicrobial defense neutralization of peptides (e.g., lysozyme, bacterial toxins defensins, ribonucleases) Cell-mediated immunity Complement proteins, CD4 T cells produce cytokines, acute-phase cytokines that induce reactants inflammation; cytotoxic Recognition of PAMPs by T lymphocytes attack PRRs such as TLRs host cells that contain intracellular bacteria 4  6/27/2024 BACTERIAL EVASION MECHANISMS A – Inhibiting chemotaxis B – Blocking adherence of phagocytes to the bacterial cells C – Blocking digestion D – Inhibiting complement c3b binding E – Cleaving IgA LABORATORY DETECTION OF BACTERIAL INFECTIONS Culture of the causative agent Grow on broth or solid media Major means of diagnosis, but may take time or may not be possible Microscopic examination Gram stain or special stains Detection of bacterial antigens Rapid testing by ELISA, LFA, or LA 5  6/27/2024 LABORATORY DETECTION OF BACTERIAL INFECTIONS (CONTINUED) Molecular detection of Can obtain results in a bacterial few hours with PCR DNA or RNA Analysis of proteins Proteomics produced by specific bacteria LABORATORY DETECTION OF BACTERIAL INFECTIONS 6  6/27/2024 LABORATORY DETECTION OF BACTERIAL INFECTIONS Serology Detects antibodies to bacterial antigens Uses: To detect and confirm infections for which other laboratory methods are not available To diagnose infections for which clinical symptoms are nonspecific Current infection indicated by presence of IgM, a high IgG titer, or a fourfold rise in antibody titer between acute and convalescent samples To determine a past exposure to an organism (IgM–, IgG+) To assess reactivation or re-exposure Disadvantages: Delay between start of infection and production of antibodies Low antibody production by immunosuppressed patients STREPTOCOCCI (GAS) Streptococci are gram-positive spherical, ovoid, or lancet-shaped organisms that are catalase negative and are often seen in pairs or chains. Transmitted person to person They are divided into groups or serotypes based on certain cell wall components, including two major proteins known as M and T proteins. M Protein 7  6/27/2024 STREPTOCOCCI Interior to the protein layer is the group-specific carbohydrate that divides streptococci into 20 defined groups, designated A–H and K–V. These are known as the Lancefield groups. Some strains possess a hyaluronic acid capsule outside the cell wall that contributes to the bacterium’s antiphagocytic properties. STREPTOCOCCI Antibodies are produced to the following exoantigens or exotoxins: Detection of host streptolysin O, deoxyribonuclease B (DNase B), antibodies to these hyaluronidase, exotoxins is important in nicotinamide adenine the diagnosis of sequelae dinucleotidase (NADase), Additional virulence such as glomerulonephritis streptokinase. factors include and acute rheumatic fever. exoantigens or exotoxins, proteins excreted by the bacterial cells as they metabolize during the course of streptococcal infections. 8  6/27/2024 CLINICAL MANIFESTATIONS OF ACUTE GROUP A STREPTOCOCCAL INFECTION Suppurative or pus forming Pharyngitis (“strep throat”) Pyoderma (impetigo) Non-suppurative or non-pus forming Scarlet fever Toxic shock syndrome Necrotizing fasciitis Treated with antibiotics GROUP A STREPTOCOCCAL SEQUELAE Acute rheumatic fever Develops 1 to 3 weeks after pharyngitis or tonsillitis in 2% to 3% of infected individuals Symptoms: fever, joint pain, inflammation of the heart Most likely caused by immune responses to streptococcal antigens that cross-react with human heart tissue Poststreptococcal glomerulonephritis May follow strep infection of the skin or pharynx Damages glomeruli, producing hematuria, proteinuria, edema, hypertension, malaise, backache, abdominal discomfort, and impairment in renal function Deposits of immune complexes containing streptococcal antigens in glomeruli 9  6/27/2024 LABORATORY DIAGNOSIS OF ACUTE GROUP A STREPTOCOCCAL INFECTIONS (SUPPURATIVE) Culture on sheep blood agar Small translucent colonies surrounded by clear zone of beta hemolysis Rapid assays to detect group A streptococcal antigens Strep Screens LFA SEROLOGIC DETECTION OF GROUP A STREPTOCOCCAL SEQUELAE The classic hemolytic method for determining the ASO titer was Antistreptolysin the first test developed to measure streptococcal antibodies. This test was based on the ability of patient antibodies to O (ASO) neutralize the hemolytic activity of streptolysin O. An ASO titer greater than 166 Todd units (or >200 IU) is considered a positive test. Tubes 1-5 no hemolysis Tube 5 trace hemolysis Remaining tubes hemolysis Tube 4 represents end point Tube 4 contains 0.8 ml of serum 1 2 3 4 5 6 7 8 9 10 diluted 100 times 12 50 100 125 166 250 333 600 625 833 The reciprocal of 0.8 is 10/8 = 1.25 1.25 x 100 yields value of 125 Todd Todd Units units. 10  6/27/2024 SEROLOGIC DETECTION OF GROUP A STREPTOCOCCAL SEQUELAE (NON-SUPPURATIVE) Antistreptolysin O (ASO) Nephelometric methods currently used that measure light scatter produced by immune complexes containing streptolysin antigen Titer elevated in 85% of patients with acute rheumatic fever Does not increase in patients with skin infection SEROLOGIC DETECTION OF GROUP A STREPTOCOCCAL SEQUELAE (NON-SUPPURATIVE) Anti-DNase B Produced by both rheumatic fever and impetigo patients Tested by EIA and nephelometric methods Streptozyme test Detects antibodies to five streptococcal products: ASO Anti-hyaluronidase (AHase) Anti-streptokinase (ASKase) Anti-nicotinamide-adenine dinucleotide (anti-NAD) Anti-DNase B 11  6/27/2024 HELICOBACTER PYLORI Gram-negative microaerophilic spiral bacterium Transmission likely by fecal-oral route Major cause of gastric and duodenal ulcers Can survive in acid environment of stomach because of production of urease, which provides a buffering zone around the bacteria Treatment with antibiotics and anti-ulcer medications If untreated, can lead to gastric carcinoma or mucosa-associated lymphoid tumors DETECTION OF HELICOBACTER PYLORI INFECTION Detect urease in stomach biopsy (CLOtest) Urea breath test H pylori antigens H pylori antibodies ELISA is method of choice IgG in serum indicates an active infection Titers decrease after successful treatment 12  6/27/2024 MYCOPLASMA PNEUMONIAE Tiny bacteria that lack a cell wall Leading cause of respiratory infections Fever, headache, malaise, and cough “Walking pneumonia” Raynaud syndrome Causes Stevens-Johnson syndrome in minority of cases Spread by respiratory droplets Culture Produces mulberry colonies with a “fried egg” appearance on specialized media LABORATORY Is gold standard but rarely performed in clinical laboratories because organism is difficult to grow DIAGNOSIS OF Antibodies to M pneumoniae M. PNEUMONIAE Most useful diagnostic assay IgM antibodies = recent infection INFECTION IgG antibodies = possible reinfection Cold agglutinins Present in about 50% of patients with M pneumoniae but not specific for the infection Cause RBC agglutination at 4°C; reversible at 37°C Molecular methods Film array respiratory panel 13  6/27/2024 RICKETTSIAL INFECTIONS Obligate intracellular gram-negative bacteria Spotted fever group (SPF) e.g., Rocky Mountain spotted fever Typhus group (TG) e.g., epidemic typhus Organisms transmitted by arthropods (ticks, mites, lice, or fleas) through biting after feeding on an infected animal Review Table 20-1 ROCKY MOUNTAIN SPOTTED FEVER (RMSF) Caused by R rickettsii Transmitted by three species of ticks Symptoms include headache, nausea, vomiting, diarrhea, skin rash; death Diagnosis Clinical presentation Serology by IFA 14  6/27/2024 SUMMARY Host–microbe relationships can be symbiotic, commensalistic, mutualistic, or parasitic Bacterial virulence factors increase an organism’s ability to cause disease; these include some bacterial structural components (endotoxin, pili, flagella, capsule) and exotoxins Endotoxin is a component that is released from the cell walls of dying gram-negative bacteria, which can cause massive cytokine production and death; exotoxins are potent toxic proteins that are released from live bacteria SUMMARY Host Innate defenses such as barriers provided by skin and defenses mucous membranes, anti-microbial peptides, C’, cytokines, against APRs, and PRR recognition bacteria Adaptive defenses involving humoral and cell-mediated include: immune responses Laboratory Culture detection Staining and microscopic observation of bacterial Rapid detection of bacterial antigens infections Molecular detection of bacterial nucleic acid can involve: Serologic detection of antibodies to bacterial antigens 15  6/27/2024 SUMMARY Group A streptococci are gram-positive bacteria that most commonly cause acute infections of the upper respiratory tract and skin; some people who are untreated can develop one of two sequelae: acute rheumatic fever or glomerulonephritis Laboratory diagnosis of acute streptococcal infections involves culture on sheep blood agar and rapid assays to detect streptococcal antigens Diagnosis of streptococcal sequelae requires serologic methods to detect antibodies to streptococcal antigens, including ASO and anti-DNase B, because the bacteria are not likely to be present when symptoms appear The streptozyme test is a rapid slide agglutination test that detects antibodies to five streptococcal products. SUMMARY Helicobacter pylori is a gram-negative, urease-producing bacterium that causes gastric and duodenal ulcers; untreated infections can progress to gastric carcinoma or MALT tumors H pylori infection can be diagnosed by urease detection in stomach biopsy tissue, the urea breath test, or an ELISA to detect H pylori antibodies; serologic tests and antigen tests of stool samples can be used to determine if the bacteria have been eliminated after treatment Mycoplasma pneumoniae are tiny bacteria that lack a cell wall; they are a major cause of respiratory infections such as “walking pneumonia” 16  6/27/2024 SUMMARY M pneumoniae is difficult to grow in culture. The main methods of detection are serologic assays for antibodies to the organism and PCR to detect DNA from the bacteria; cold agglutinins are produced in about one-half of patients Rickettsia are obligate intracellular gram-negative bacteria that are transmitted by arthropods; two diseases caused by rickettsia are RMSF and typhus. Serologic testing for antibodies to R rickettsii by IFA is considered the gold standard for laboratory diagnosis of RMSF POSTAMBLE READ the TEXTBOOK for the details to answer the UNIT OBJECTIVES. USE THE UNIT OBJECTIVES AS A STUDY GUIDE All test questions come from detailed material found in the TEXTBOOK (Not this PowerPoint) and relate back to the Unit Objectives 17