NonfermentersNonfermenters Gram-Negative Bacilli

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

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

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

Morphology and Characteristics Gram(-) bacilli, coccobacilli Obligate aerobes Most will not grow or grow poorly under anaerobic conditions Some require hours for growth Grow best at 37 0 C, but a few grow better at RT (Ps. fluorescens) or tolerate higher (42ºC, Ps. aeruginosa)

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

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

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 42 0 C

Nonfermenters: Lab Unknowns Oxidase O/F Motile SXT NO 3 Red Pseudomonas + +/- + R + Acinetobacter - +/- - S - Stenotrophomonas - -/- + S - Chyseobacterium + +/- - R -

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

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 –LD 50 in mice= ng –Blocks host cell protein synthesis –Liver is prime target

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)

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

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

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

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

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

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)

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).

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

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

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)

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

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

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

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

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

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.

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.

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?