Nonfermenters

1. Nonfermenters Gram-Negative Bacilli

2. Clinically Important Aerobic Gram(-) Bacilli • 75% = Facultative anaerobic fermenters - Enterobacteriaceae • 15% = Aerobic nonfermenters – Pseudomonadaceae and related bacteria • 10% = Pasteurellaceae • <1% = Unusual bacilli

3. Nonfermenters: Gram(-) Bacilli • Inhabit soil, vegetation, water; harmless parasites on mucous membranes of human and animals • Simple growth requirements • Hospital moist reservoirs – food, ice machine, cut flowers, sinks, toilets, floor mops, disinfectant solutions, respiratory therapy equipment • Numerous virulence factors • Broad antimicrobial resistance • Infections primarily opportunistic – colonize, infect immunocompromised; gain access to normally sterile body site through trauma

4. Nonfermenters: Genera • No family designation • Many genera whose names continually changing • Do not ferment glucose • Pseudomonas aeroginosa • Acinetobacter baumannii • Stenotrophomonas maltophilia (former Ps.) • Burkholderia cepacia (former Ps.) • Moraxella catarrhalis

5. Morphology and Characteristics • Gram(-) bacilli, coccobacilli • Obligate aerobes • Most will not grow or grow poorly under anaerobic conditions • Some require 48-72 hours for growth • Grow best at 370 C, but a few grow better at RT (Ps. fluorescens) or tolerate higher (42ºC, Ps. aeruginosa)

6. Lab Culture Media • Nonfastidious, isolated same as Enterobacteriaceae • Mac plate – some grow • CBA, MH plate - morphology, size, hemolytic activity, pigmentation (green, blue) provide valuable information for ID

7. Lab Test ID • TSI= K/K - No glucose fermentation • Oxidase (±) • Mac plate (±) Growth • Unusually resistant to antibiotics

8. CDC Scheme Nonfermenters (8 Groups) • Mac plate - Growth, no growth • Oxidase • O/F carbohydrates • Further testing: • Motility • Nitrate reduction • Urease production • Esculin hydrolysis • Indole production • Rapid amino acid decarboxylation • Pigment production • Phenylalanine deaminase • Growth at 420 C

9. Nonfermenters: Lab Unknowns Oxidase O/F Motile SXT NO3Red Pseudomonas + +/- + R + Acinetobacter - +/- - S - Stenotrophomonas - -/- + S - Chyseobacterium + +/- -R -

10. Pseudomonas aeruginosa • “false unit”– in pairs, resemble single cell • Numerous virulence factors • Broad-based antimicrobial resistance • Some strains mucoid (CHO capsule), common in cystic fibrosis patients • Diffusible pigments – pyoverdin (fluorescein, yellow), pyocyanin (blue), pyorubin (red) • Tolerate temperatures (4º-42ºC) • Sweet grapelike odor on culture plate

11. Ps. aeruginosa:Virulence Factors (Extracellular) • Protease - • Tissue destruction • Degrade Complement, IgA • Inhibit neutrophil • Elastase – destroy elastin fiber of lung tissue, blood vessels; hemorrhagic lesions, spreading of infection • Exotoxin - most toxic product • Cytotoxin lethal for many mammals • LD50 in mice= 60-80 ng • Blocks host cell protein synthesis • Liver is prime target

12. Virulence Factors (Extracellular) • Phospholipase C – attack lipid of cell membrane • Hemolysin • Breakdown of phospahtidyl choline, a major surfactant of the lung; lead to tissue damage, pulmonary collapse • Leukocidin – cytotoxic • Pyocyanin – secreted pigment • Toxic • Generate reactive oxygen intermediates (superoxide radical, hydrogen peroxide)

13. Ps. aeruginosa:Virulence Factors (Cell Surface) • Pili and non-pilus adhesions – attachment respiratory epithelium • LPS – endotoxin; sepsis syndrome, DIC • Iron capturing ability – nutrition, growth • Flagella – motility • Alginate synthesis – forms viscous gel around MO, function as adhesion, also prevent phagocytosis • Outer membrane changes - antibiotic unable to enter bacterial cell; drug resistance

14. Ps. aeruginosa:Respiratory Tract Infection • Leading cause nosocomial RTI • Range from colonization, benign tracheobronchitis to severe necrotizing bronchopneumonia • Seen in patients with: • Cystic fibrosis • Chronic lung diseases • Neutropenia • Frequently following use of contaminated respiratory therapy equipment • Severe infections lead to bacteremia and higher mortality

15. Ps. aeruginosa:Bacteremia, Endocarditis • Higher mortality rate due to: • Virulence of Pseudomonas strain • Infection in immunocompromised (neutropenia patient, diabetes mellitis, extensive burns, hematologic cancers) • Originate from initial infections of LRT, UT, skin & soft tissue (burns, wounds) • Endocarditis commonly seen in IV drug abusers

16. Ps. aeruginosa:Ear Infection • External otitis media: • Swimming a risk factor (swimmer’s ear) • Manage with topical antibiotics, drying agents • Malignant external otitis media: • More virulent form, invade underlying tissue • Can be life threatening • Require aggressive antimicrobials + surgery • Chronic otitis media

17. Ps. aeruginosa:Burn Infection • Colonize burn wound • Localize vascular damage, tissue necrosis, bacteremia • Factors predispose patient to infection: • Moist surface of burn • Absence of neutrophil response • Limited success treating with topical creams and wound management

18. Ps. aeruginosa: Other Infections • GI, UT, CNS, eye, musculoskeletal • Underlying conditions: • Presence of Pseudomonas in a moist reservoir • Circumvention or absence of host defense (e.g. cutaneous trauma, elimination of NF by injudicious use of antibiotics, neutropenia) • Indwelling urinary catheter (best to remove ASAP)

19. Pseudomonas: Treatment • Typically resistant to most antibiotics • Difficult to treat patient - often with compromised host defense, unable to augment antibiotic activity • Important to isolate MO for antibiotic susceptibility testing • Requires combined treatment: • Aminoglycoside (tobramycin) • β-lactam antibiotic (ceftaidine, piperacillin).

20. Pseudomonas: Prevention • Hospital Infection Control • Avoid contamination of sterile equipment such as respiratory therapy machine • Prevent cross-contamination of patient by medical personnel; i.e. hand washing, fomites • Avoid inappropriate use of broad-spectrum antibiotics that kill and suppress host NF

21. Acinetobacter • “unable to move” • A. baumannii(oxidizer = saccharolytic) • A. lwoffii(nonoxidizer = inert) • Found in soil, water; NF skin, oropharyngeal • Common colonizer, cause of nosocomial respiratory infection • Thrive in moist environments, found as contaminants in respiratory equipment and monitoring devices • Resistant to many antibiotics; use aminoglycosides and broad-spectrum cephalosporins

22. Stenotrophomonas maltophilia • “narrow, feed, unit”; “malt lover” • Second most frequently isolated nonfermenter • Nosocomial - transient NF of patients • Opportunist – especially debilitated patient, impaired host-defense • Variety of infections – bacteremia, pneumonia, meningitis, wounds, UTIs • Resistant to commonly used antibiotics • Treat with trimethoprim-sulfmethaxazole (SXT)

23. Burkholderia cepacia • “onion” • Low level virulence, nosocomial pathogen • Respiratory Tract infection: • Range from colonization to broncopneumonia • Patients with cystic fibrosis, chronic granulomatous disease • Opportunistic infection: • Patient with urinary catheter • Immunocompromised patient with intravascular catheter

24. Burkholderia pseudomallei:Melioidosis • Disease primarily SE Asia, India, Africa, Australia; normal inhabitant of soil, water • Acquired via contamination of wounds, inhalation, ingestion • Range of infection: • Most unapparent, asymptomatic • Cutaneous, localized suppurative infection, lymphadenopathy, fever, malaise; resolves or progress to sepsis • Chronic or acute pulmonary infection, overwhelming septicemia with multiple abscesses in many organs

25. Burkholderia mallei: Glanders • “mallei” glanders • Equine infection, humans occasionally acquire disease • Contact with infected nasal secretions of horses; through skin abrasions, occasionally inhalation • A problem in military when horses were commonly used • Disease may manifest as: • Chronic pulmonary disease • Multiple abscesses of skin, subcutaneous tissue, lymphatics • Acute, fatal septicemia • Potential bioterrorist agent

26. Moraxella catarrhalis • “downflowing, inflammation” • Oropharyngeal NF • Previously healthy patient, also hospitalized patient • Bronchitis, bronchopneumonia in patient with chronic pulmonary disease • Sinusitis • Otitis media • Most penicillin-resistant, susceptible to erythromycin

27. Class Assignment • Textbook Reading: Chapter 21 Nonfermenting Gram-Negative Bacilli (Omit: Less Commonly Encountered Nonfermentive GNB) • Key Terms • Learning Assessment Questions

28. Case Study 4 – Pseudomonas • A 63-year-old man has been hospitalized for 21 days for the management of newly diagnosed leukemia. • Three days after the patient entered the hospital, a urinary tract infection with Escherichia coli developed. • He was treated for 14 days with broad-spectrum antibiotics. • On day 21 of his hospital stay the patient experienced fever and shaking chills.

29. Case Study 4 - Pseudomonas • Within 24 hours he became hypotensive, and ecthymic skin lesions appeared. • Despite aggressive therapy with antibiotics, the patient died. • Multiple blood cultures were positive for P. aeruginosa.

30. Case Study - Questions • 1. What factors put this man at increased risk for infection with P. aeruginosa? • 2. What virulence factors possessed by the organism make it a particularly serious pathogen? What are the biologic effects of these factors? • 3. What antibiotics can be used to treat P. aeruginosa? • 4. What diseases are caused by S. maltophila? A. baumanni? M. catarrhalis?