Haemophilus and Other Fastidious Bacteria 2007-2011 PDF

Haemophilus and Other Fastidious Gram-Negative Bacilli Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 1  General Characteristics ⬤ Family Pasteurellaceae Haemophilus & Pasteurella Gram-negative coccoid to rod-shaped...

Haemophilus and Other Fastidious Gram-Negative Bacilli Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 1  General Characteristics ⬤ Family Pasteurellaceae Haemophilus & Pasteurella Gram-negative coccoid to rod-shaped Nonmotile Aerobic to facultatively anaerobic Convert nitrate to nitrate Oxidase + and catalase + Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 2  Gram Stain of Hemophilus 3 S.A. Basit  Haemophilus Species ⬤ Gram-negative pleomorphic coccobacilli or rods Coccobacilli in direct smears Rods with occasionally long filamentous rods from colony growth ⬤ Biochemical reactions Nonmotile, oxidase +, catalase +, nitrate reduction Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 4  Haemophilus Species (Cont’d) ⬤ Obligate parasites of the mucous membranes ⬤ 10 species involved in human infection Emphasis on H. influenzae, H. haemolyticus, H. aegyptius, H. influenzae biotype aegyptius, and H. ducreyi Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 5  Haemophilus Species (Cont’d) ⬤ Haemophilus – Blood loving Require X and V factors X factor is hemin, hematin V factor is nicotinamide-adenine dinucleotide (NAD) ⬤ Para – require only V factor Haemophilus parainfluenzae Produces X factor requires V factor Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 6  Haemophilus Species (Cont’d) ⬤ Hemolysis of 5% horse or rabbit blood agar H. hemolyticus species and sometimes H. ducreyi Sheep blood agar only contains X factor and not V factor Thus, some will not grow without V factor added because NADases are present in BAP destroy V factor Chocolate agar release X and V factor and deactivates NADases Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 7  Growth Patterns ⬤ Satellitism Growth of a fastidious organisms around other bacteria that release the necessary growth factors or breakdown toxic products S. aureus, S. pneumoniae, Neisseria species that cause hemolysis release the factors or naturally produce V factor Exceptions H. aphrophilus and H. ducreyi Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 8  Satellitism 9 S.A. Basit Example of Satellitism Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 10  Growth Patterns (Cont’d) ⬤ 10% of normal flora of the upper respiratory tract in adults 2%-6% in children from birth through childhood with a higher percentage colonization in daycare centers As they mature they convert from encapsulated to nonencapsulated strains Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 11  Haemophilus influenzae: Historical Perspective ⬤ H. influenzae was isolated from pandemic influenza victims Wasn’t cause of disease Was frequently isolated Secondary infections Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 12  Virulence Factors ⬤ Capsule Serotypes a-f ⬤ IgA protease Cleaves IgA on mucous membranes ⬤ Outer membrane proteins and lipopolysaccharide (LPS) Not well defined Antibody to these proteins is somewhat protective LPS paralyzes the cilia Can’t clear the lungs ⬤ Adherence Pili and other structures Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 13  Virulence Factors (Cont’d) ⬤ Outer membrane proteins and lipopolysaccharide (LPS) Not well defined Antibody to these proteins is somewhat protective LPS paralyzes the cilia Can’t clear the lungs ⬤ Adherence Pili and other structures Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 14  Capsule ⬤ Classification based on capsule serotype Contains ribose, ribitol, and phosphate Antiphagocytic and anticomplement activity ⬤ Haemophilus influenzae serotype b (Hib) Primarily cause disease in children Causes bacteremia and spread to other tissues Hib vaccine is useful in reducing incidence of disease Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 15  Capsule (Cont’d) ⬤ Nontypable strains (no capsule) Invade the respiratory tract and tissues located around the same area Localized infections Pneumonia, otitis media, sinusitis Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 16 17 S.A. Basit 18 S.A. Basit 19 S.A. Basit  Types of Hemophilus Infection 20 S.A. Basit  Clinical Manifestations of Haemophilus influenzae ⬤ Diseases of Hib strains Meningitis Invasion of the respiratory tract Bacteremia allows spread to the meninges and CSF Infants and children 3 months to 6 years old ⬤ Epiglottitis Acute inflammation and swelling causing airway obstruction Affects children 2 to 4 years old Emergency tracheostomy Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 21  Clinical Manifestations of Haemophilus influenzae (Cont’d) ⬤ Bacterial tracheitis Life-threatening disease in young children ⬤ Cellulitis Cheek causing swelling, pain, and reddish blue color of the inflamed area Children younger than 2 years old Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 22  Clinical Manifestations of Haemophilus influenzae (Cont’d) ⬤ Other infections Acute pharyngitis and pneumonia Children about 1 year old ⬤ Diseases of nonencapsulated strains Otitis media, bronchitis, sinusitis, pneumonia in the elderly, genital tract infections Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 23 Infections caused by H. influenzae Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 24  Haemophilus aegyptius ⬤ H. aegyptius and H. influenzae biogroup aegyptius can cause conjunctivitis Pinkeye Acute, contagious, purulent conjunctivitis Particularly in children ⬤ H. influenzae biogroup aegyptius Brazilian purpuric fever (BPF) Occurs in warm tropical climates Recurrent conjunctivitis, high fever, vomiting, petechiae, purpura, septicemia, and shock High mortality as high as 70% Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 25  Haemophilus ducreyi ⬤ Sexually transmitted infection Chancroid ⬤ Incubates 4-14 days Painful lesion with an irregular edge Genital and perianal areas Penis Labia or vagina Enlarged and draining lymph nodes Buboes Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 26  Hemophilus ducreyi Chanchoid bacilli, don’t demonstrate “Satellite Phenomenon” Soft chance: red papule (hungry hyperemia) → pustule, ulcer → lymphadenopathy →bubo 27 S.A. Basit  Chancroid Lesions Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 28  H. ducreyi ⬤ Dmelco’s test and Ducreyi’s: hypersensitivity reaction tests ⬤ MOT: STD, direct contact with lesions ⬤ Coccobacilli “School of fish” ⬤ CAP supplemented with 1% IsoVitalex 29 S.A. Basit Miscellaneous Haemophilus Species ⬤ H. parainfluenzae Endocarditis ⬤ H. aphrophilus Bite wound infections Endocarditis Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 30  Hemophilus parainfluenzae Normal inhabitant of respiratory tract Zoonotic disease of swine ”hog influenza” Normal flora of nasopharynx and URT Normal Flora → also cause bacterial endocarditis and pneumonia Found in female GUT associated with vaginitis 31 S.A. Basit  Specimen Processing ⬤ Haemophilus species Die rapidly plated within 10 minutes for maximum recovery H. ducreyi Clean specimen site with sterile saline and sterile gauze Swab base of ulcer with cotton swab moistened with sterile saline ⬤ H. influenzae Chocolate agar w/bacitracin Helps inhibit other organisms especially respiratory flora Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 32  Specimen Processing (Cont’d) ⬤ H. aegyptius Chocolate supplemented with 1% isovitaleX ⬤ H. ducreyi GC agar (hemoglobin, FBS, vancomycin to reduce genital flora) ⬤ Grow in a capnophilic atmosphere ⬤ Humidity chamber Prevent drying Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 33 Direct Smear of H. influenzae Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 34  Culturing Haemophilus ⬤ Besides chocolate agar BAP for groups that don’t require V factor H. aphrophilus MacConkey agar ⬤ Colony morphology Translucent, moist, smooth, convex colonies Mousy or bleach like odor Encapsulated strains appear more mucoid Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 35  Culturing Haemophilus (Cont’d) ⬤ H. influenzae Gram – coccobacilli or small rods ⬤ H. ducreyi Gram – coccobacilli arranged in groups “school of fish” Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 36 Colony and Gram Stain of H. influenzae Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 37 Colony and Gram Stain of H. influenzae (Cont’d) Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 38  Laboratory Identification ⬤ Take colonies from an initial isolation of Haemophilus Do not pick up the growth media ⬤ Place the colonies in nutrient broth and mix ⬤ Plate inoculated nutrient broth on a nutrient agar plate or a Mueller-Hinton agar plate Add X and V strips to the media Incubate at 35-37°C in 5-10% CO2 18-24 hrs ⬤ Read plates Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 39 Organism Requiring X and V Factors Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 40 Organism Requiring V Factor Only Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 41 Organism Requiring X Factor Only Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 42  Porphyrin Test ⬤ If they don’t require X factor Porphyrin + (make X) ⬤ Delta-aminolevulinic acid (ALA) gets converted to Porphyrins or Protoporphyrins Kovacs reagent Red color produced in lower part of the tube Wood UV lamp fluoresce reddish orange under UV light Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 43 Example of Porphyrin Test Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 44 Differential Tests for Haemophilus Species Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 45 Differential Tests for Haemophilus Species (Cont’d) Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 46  Treatment ⬤ Cefotaxime or ceftriaxone Recommended ⬤ Trimethoprim sulfamethoxazole, imipenem, ciprofloxacin, and chloramphenicol Alternative treatments ⬤ H. ducreyi Erythromycin Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 47  HACEK ⬤ H – Haemophilus spp. Now Aggregatibacter aphrophilus ⬤ A – Actinobacillus actinomycetemcomitans Now Aggregatibacter actinomycetemcomitans ⬤ C – Cardiobacterium hominis ⬤ E – Eikenella corrodens ⬤ K – Kingella spp. Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 48  HACEK Group General Characteristics ⬤ Gram-negative bacilli ⬤ Require an increased CO2 (5%-10%) environment ⬤ Significant cause of endocarditis ⬤ Usual flora of the oral cavity Thus are pathogens associated with human bite wounds causing septicemia and subacute endocarditis ⬤ Opportunists in immunocompromised hosts Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 49 Differential Tests for Haemophilus Biogroups Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 50  A. aphrophilus ⬤ Aggregatibacter aphrophilus From the Greek word “foam loving” Described earlier Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 51 A. aphrophilus (Cont’d) Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 52 A. aphrophilus (Cont’d) Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 53 A. aphrophilus (Cont’d) Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 54  Aggregatibacter actinomycetemcomitans ⬤ Primarily an animal pathogen but found in human oral cavity Nonmotile gram-negative coccobacilli 24-48 hours to grow Distinctive star formation in the center of the colonies Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 55  Aggregatibacter actinomycetemcomitans (Cont’d) ⬤ Ferments carbohydrates in the presence of serum Not lactose, or sucrose ⬤ Catalase + oxidase variable ⬤ No growth on Mac plates ⬤ Urease, indole, esculin, citrate negative Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 56 Photographs of Actinobacillus actinomycetemcomitans Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 57  Photographs of Actinobacillus actinomycetemcomitans (Cont’d) Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 58  Cardiobacterium hominis ⬤ Pleomorphic, nonmotile, gram – bacillus Sometimes appear false positive on Gram stain Show “rosette” morphology ⬤ Normal flora of the nose, mouth, and throat Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 59 Cardiobacterium hominis (Cont’d) ⬤ Involved in endocarditis Grows on the heart valve and is resistant to antibiotics so valve must be replaced ⬤ Grow on BAP, chocolate agar Not Mac plates ⬤ Pitting may be produced on agar plates Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 60 Photographs of Cardiobacterium hominis Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 61 Photographs of Cardiobacterium hominis (Cont’d) Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 62  Eikenella corrodens ⬤ Normal flora of oral and bowel cavities Fight and bite wounds ⬤ Infections Meningitis Pneumonia Osteomyelitis Arthritis Post-op tissue infections usually from bacteremia associated with wounds Cellulitis Inoculation with needles “licked clean” Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 63  Eikenella corrodens (Cont’d) ⬤ Grow in capnophilic environment ⬤ Require hemin (X factor) ⬤ Gram-negative coccobacilli Bleachlike odor with yellow colonies pitting the agar (corrodens) ⬤ Biochemical tests ⬤ Nonmotile, nonfermenting, oxidase +, catalase –, lysine decarboxylase +, ornithine decarboxylase +, arginine decarboxylation – Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 64  Eikenella corrodens (Cont’d) ⬤ Biochemical tests Nonmotile Nonfermenter Oxidase + and catalase – Lysine decarboxylase + Ornithine decarboxylase + Arginine decarboxylation – Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 65 Photographs of Eikenella corrodens Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 66 Photographs of Eikenella corrodens (Cont’d) Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 67  Kingella Species ⬤ Coccobacilli or short rods in pairs or chains ⬤ Nonmotile but may twitch ⬤ Oxidase + catalase – sugar fermenters Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 68  Kingella Species ⬤ Kingella dentrificans May grow on Thayer Martain medium and resemble N. gonorrhoeae colonies do not pit Gram-negative rods have square ends ⬤ Biochemical tests Glucose + Reduces nitrate (+) Urease – Indole – Esculin – Gelatin – Citrate – Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 69  Kingella Species (Cont’d) ⬤ Kingella kingae Isolates of children less than 5 years old Negative for sucrose Positive for glucose, maltose Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 70 Kingella Species (Cont’d) Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 71  Capnocytophaga ⬤ Previously called DF-1 and DF-2 (Dysgonic fermenters) ⬤ Gram-negative bacilli often fusiform May resemble Fusobacterium ⬤ Facultatively anaerobic Require CO2 ⬤ More common in septicemia than endocarditis Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 72 Photographs of Capnocytophaga Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 73 Photographs of Capnocytophaga (Cont’d) Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 74 Photographs of Capnocytophaga (Cont’d) Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 75 76 S.A. Basit 77 S.A. Basit  Pasteurella multocida ⬤ Contains five serogroups A-F ⬤ Gram-negative pleomorphic coccobacilli with bipolar staining Oval shaped, short rods, longer filaments ⬤ Grow on BAP and chocolate agar Nonhemolytic mucoid colonies with a narrow green or brown halo Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 78 79 S.A. Basit  Pasteurella multocida (Cont’d) ⬤ Colonize the URT and GI tract of mammals and birds Exposure from dog and cat bites or scratches ⬤ Biochemical tests Nonmotile Catalase + Oxidase + Glucose fermentation Weak acid (sick acid) production Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 80 Differential Characteristics of Pasteurella Species Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 81 Plate Growth of Pasteurella multocida Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 82  Brucella Species ⬤ Brucellosis or undulant fever Reoccurring fever at regular intervals that persist for days, months, or years ⬤ Zoonotic infection acquired from animals or animal products Organism found in urine and milk ⬤ slow growing fastidious organisms ⬤ can grow in BAP and occasionally in MAC ⬤ treatment: tetracycline in combination with spectinomycin and gentamicin Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 83  Brucella Species (Cont’d) ⬤ Most common species for human brucellosis B. melitensis (most common isolate), B. abortus, B. canis, B. suis ⬤ Can produce abortion secondary to bacteremia in pregnant women Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 84  Human Brucellosis ⬤ portal of entry: intestinal tract, mucous membrane and skin ⬤ malta fever; undulant fever ⬤ Bang’s bacillus 85 S.A. Basit  Species of Brucella Brucella Brucella abortus melitensis Brucella suis 86 S.A. Basit Brucella canis  Symptoms of Human Brucellosis 1. B. melitensis - severe acute disease with complications 2. B. abortus - mild disease with suppurative complications, uncommon 3. B. suis - suppurative, destructive disease with chronic manifestations 4. B. canis - mild disease with suppurative complications, uncommon 87 S.A. Basit 88 S.A. Basit  Characteristics of Brucella Species ⬤ Strict aerobes (some require CO2) ⬤ Nonspore forming gram-negative coccobacilli ⬤ Nonmotile, nonencapsulated, intracellular parasites Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 89  Characteristics of Brucella Species (Cont’d) ⬤ Biochemical tests Oxidase + Catalase + H2S production and urease help differentiate ⬤ Blood and bone marrow cultures Acute and convalescent sera Category B select biologic agent Serology test are safer Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 90 Colonies of Brucella melitensis Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 91 Differential Characteristics of Brucella Species Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 92 Differential Characteristics for ID of Brucella from Similar Organisms Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 93  Francisella Species ⬤ Facultative intracellular parasite ⬤ Nonmotile, small gram-negative coccobacilli ⬤ Strict aerobe ⬤ Zoonotic infection Causes tularemia Acute febrile, granulomatous disease with rapid onset and flulike symptoms Category A select agent Diagnosis from serology Culture is less safe Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 94  Francisella Species ⬤ Four subspecies F. tularensis subsp. tularensis (type A)- more virulent Transmitted by rabbits, sheep, ticks F. tularensis subsp. holarctica (type B) Transmitted by rodents and mosquito F. tularensis subsp. mediasiatica F. tularensis subsp. Novicida Similar to F. tularensis but less lethal Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 95  Francisella tularensis Gram (-) aerobic bacillus Tularemia :grandular fever, rabbit fever, deerfly fever, tick fever. (is frequently spread by direct contact with rabbits, the disease can also be spread by other animals, typically rodents, and by arthropods. ) Edward Francis: Tulare County, California Worldwide distribution: 2 biochemical varieties 96 S.A. Basit 97 S.A. Basit 98 S.A. Basit  2 major types 99 S.A. Basit Infected individuals are normally placed on a regimen of streptomycin or gentamycin for 10-14 days 100 S.A. Basit  Clinical Manifestations 101 S.A. Basit Colony Growth on Chocolate Agar for F. tularensis subsp. tularensis Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 102  Legionella Species ⬤ Legionella pneumophila Legionnaires’ disease Pontiac fever 14 serogroups ⬤ Sources Present in environmental water sources Air conditioning systems Respiratory secretions on objects used by patient Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 103  Legionella Species (Cont’d) ⬤ Infection Disease has a large range from asymptomatic to deadly Results from the inhalation of aerosols Note: Person to person via aerosol transmission has not been documented ⬤ Detection Culture Antigen in the urine Direct fluorescent antibody staining Serology Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 104  Legionella pneumophila motile, rod-shaped, gram(-), aerobic, non-capsulated, non-fermentative This disease was first discovered in 1976 among a group of elderly men attending an American Legion Convention in Philadelphia, 34 out 221 persons died. 105 S.A. Basit  Legionella Species ⬤ Legionella pneumophila Legionnaires’ disease Pontiac fever 14 serogroups ⬤ Sources Present in environmental water sources Air conditioning systems Respiratory secretions on objects used by patient Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 106  Legionella Species (Cont’d) ⬤ Infection Disease has a large range from asymptomatic to deadly Results from the inhalation of aerosols Note: Person to person via aerosol transmission has not been documented ⬤ Detection Culture Antigen in the urine Direct fluorescent antibody staining Serology Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 107 108 S.A. Basit 109 S.A. Basit 110 S.A. Basit 111 S.A. Basit  Legionnaires’ Disease ⬤ Sporadic cases – community acquired ⬤ Epidemic outbreaks Usually from air conditioners ⬤ Nosocomial infections in immunocompromised Respiratory equipment and aerosols ⬤ One of the top four pathogens in community acquired pneumonia Streptococcus pneumoniae, Mycoplasma pneumoniae, Chlamydia pneumoniae Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 112  Legionnaires’ Disease (Cont’d) ⬤ Incubation period is 2 to 10 days Symptoms Nonproductive cough, fever, headache, and myalgia Progressing to bloody or purulent sputum, rales (crackling), difficulty breathing, and shaking chills Dissemination via circulatory system can occur Invades the kidneys, liver, heart, CNS, and lymphatic system Destruction leads to death in pneumonia doesn’t Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 113  Legionnaires’ Disease (Cont’d) ⬤ Mortality rate from 15%-30% sometimes approaching 50% ⬤ Most cases are serogroup 1 (85%) Also serogroup 4 and 6 Sometimes L. micdadei Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 114 115 S.A. Basit  Pontiac Fever ⬤ Incubation period of 2 days Flulike symptoms Fever, headache, myalgia Lasting 2-5 days ⬤ Spontaneous recovery Doesn’t generally lead to mortality Mild form of Legionella Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 116  Reservoirs of Legionella ⬤ Most aquatic sources Lake, river, springs, man-made water treatment systems Can survive in chlorinated water ⬤ Specialized sources Heating and cooling towers of buildings Fountains Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 117 Reservoirs of Legionella (Cont’d) ⬤ Can colonize a variety of sources due to: Ability to multiply at a large range of temps 20-43° C Survive in temps 40 to 60° C Adherence to pipes, rubber, and plastics Can live intracellularly in protozoa Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 118  Clinical infections of Legionella ⬤ Intracellular parasite of cells Enter and multiply within host cells Particularly bronchoalveolar macrophages ⬤ Predisposition for disease Immunocompromised Patients with chronic lung disease Alcoholics and heavy smokers Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 119  Identifying Legionella ⬤ Specimens Predominantly respiratory specimens BAL, sputum, bronchial washings Urine specimens for antigen test Soluble antigen found in the urine for serogroup 1 Stays positive for a long time thus only confirms infection happened Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 120  Identifying Legionella (Cont’d) ⬤ Direct examination Pleomorphic thin weakly staining gram-negative rods Found within macrophages and neutrophils as well as extracellularly ⬤ Urine antigen test ⬤ Direct antigen tests available Fluoresce if positive for Legionella Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 121  Culturing Legionella ⬤ Aerobic bacteria ⬤ Require L-cysteine for growth so do not grow on routine media ⬤ When suspected Grow on BCYE Grow as small pinpoint colonies Acid wash specimens to reduce contaminants Dilute sample 1:10 in 0.2N KCl-HCl solution for 5 minutes Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 122  Culturing Legionella (Cont’d) ⬤ Grow plates in normal atmosphere at 35-37° C for at least 7 days ⬤ Grayish white or blue green convex, wet looking colonies Confirmation Requires proving L-cysteine is required for growth Can use molecular methods Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 123 124 S.A. Basit BCYE Agar with Legionella pneumophila Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 125 Gram Stain of Legionella pneumophila Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 126 Grouping of Legionella Species Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 127 Common Phenotypic Characteristics of Legionella Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 128 Schema for ID of Legionella Organisms Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 129 130 S.A. Basit Example of DFA Test for Legionella Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 131  Antimicrobial Susceptibility ⬤ Susceptibility not routinely performed Use erythromycin or in combination with rifampin ⬤ Alternatives Doxycycline, trimethoprim-sulfamethoxazole, newer macrolides, or fluoroquinolones Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 132  Bordetella ⬤ Bordet-Gengou bacillus ⬤ small strict aerobic Gram – bacilli ⬤ pertussis (Latin: severe cough) ⬤ whooping cough ⬤ enriched media 🡪 charcoal, starch, blood and albumin 133 S.A. Basit  Bordetella Species ⬤ Seven major species Major pathogens Bordetella pertussis Bordetella parapertussis Both of these are agents of whooping cough Bordetella bronchiseptica Opportunistic pathogen involved in pneumonia and wound infections Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 134 135 S.A. Basit Bordetella Species (Cont’d) Minor pathogens Bordetella avium Bordetella hinzii Bordetella holmesii Bordetella trematum Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 136  Infection of Bordetella pertussis and parapertussis ⬤ Breathing in aerosols Bacteria adhere to and grow on ciliated respiratory epithelial cells ⬤ Highly contagious disease 90% of exposed contacts get disease Vaccine widely used; however, effectiveness drops rapidly after administered Adults can become transient carriers With and without symptoms Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 137 138 S.A. Basit  Virulence Factors ⬤ Filamentous hemagglutinin (FHA) Facilitate attachment to epithelial cells ⬤ Pertussis toxin (PT) Interfere with signal transduction ⬤ Adenylate cyclase toxin Increases cAMP levels ⬤ Tracheal cytotoxin Ciliostasis - cilia don’t clear the airway Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 139  Clinical Infections and Disease Course ⬤ Whooping cough ⬤ Exposure and incubation period of 1-2 weeks ⬤ Initially cold or flulike symptoms, sneezing, runny nose Catarrhal phase 1-2 weeks (highly contagious) ⬤ Sudden severe repetitious coughing Followed by “whoop” Rapid gasp for air Can be followed by vomiting Paroxysmal phase Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 140  Whooping Cough Clinical Duration Symptoms Syndromes Catarrhal 1-2 Rhinorrhea, malaise, fever, weeks sneezing, anorexia Paroxysmal 2-4 Repetitive cough with whoops and weeks vomitting Convalescent 3-4 Diminished paroxysm weeks Cough 2nd complications: pneumonia, seizures and encephalopathy 141 S.A. Basit 142 S.A. Basit  Clinical Infections and Disease Course ⬤ Serious infection in young children Cyanosis from lack of oxygen Sometimes need assistance in keeping airway open ⬤ Convalescent phase Begin recovery about 4 weeks after initial symptoms Decrease in bouts of coughing but may take weeks or months for complete resolution Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 143  Diagnosis and Isolation ⬤ Specimens are best from the nasopharynx Aspirates of fluid Calcium alginate or Dacron swabs up each nasal passage as deep as possible ⬤ Inoculate a transport system Regan-Lowe transport medium Direct to plate media Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 144  Diagnosis and Isolation (Cont’d) ⬤ Culture Bordet-Gengou potato infusion agar with glycerol and sheep blood Alternative is charcoal agar with 10% horse blood and cephalexin Also plate to BAP and MacConkey agar Chocolate if you need to rule out Haemophilus ⬤ Incubate plates 35° C in aerobic conditions 7 days Adequate moisture to prevent plate drying Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 145 Summary of Laboratory Diagnosis of Pertussis by Culture Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 146  Colony Morphology ⬤ Gram-negative rods or coccobacilli ⬤ Smooth silver pinpoint colonies resembling mercury droplets ⬤ Direct fluorescent antibody tests are used for confirmation Agglutination tests Require larger amount of organism ⬤ Biochemicals to differentiate species Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 147 Colonies of Bordetella pertussis Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 148 Differential Characteristics of Bordetella Species Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 149  Antimicrobial Susceptibility ⬤ Erythromycin Most common ⬤ Azithromycin Elsevier items and derived items © 2011, 2007 by Saunders, an imprint of Elsevier Inc. 150