Gram-positive Bacilli

1. Gram-positive Bacilli Ken B, Waites, M.D. F(AAM) Professor of Pathology Director of Clinical Microbiology

2. Objectives • To review and discuss: • microbiological characteristics • epidemiology • virulence factors • associated diseases • laboratory detection of clinically important gram-positive bacilli. Organism groups to be discussed include: • Bacillus • Listeria • Erysipelothrix • Corynebacterium • Rhodococcus • Nocardia • Tropheryma

3. Bacillus • Ubiquitous in soil worldwide • Endospore-forming • Aerobic/facultative anaerobic • Gram-positive bacilli • Can be gram-variable • Often  hemolytic & motile • (except B. anthracis) • Usually catalase positive B. cereus - Gray, spreading -hemolytic colonies on SBA

4. Bacillus Endospores • Thick walled structures formed by vegetativecells • Resistant to radiation, chemicals, heat, desiccation  (dipicolinic acid) • Steam autoclaving necessary for destruction

5. Bacillus: Clinical Significance • Common environmental commensals • Occasional opportunistic pathogens & culture contaminants • B. cereus group • B. anthracis • Others

6. Bacillus cereus Group • Gastroenteritis • Heat-stable enterotoxin (emetic form) • Heat-labile enterotoxin (diarrheal form) • Similar to V. cholera – stimulates cAMP watery diarrhea • Contaminated rice, meat, vegetables • Other Clinical conditions • Ocular infections after trauma • Necrotic toxin, hemolysin, phospholipase C • IV-catheter-related sepsis, endocarditis, meningitis • Immunocompromised persons and drug abusers

7. B. anthracis: Anthrax • Rare in US due to control in animals • Enzootic in middle east • Transmitted by contact with animal products • Spores remain infectious for years • Usually cutaneous inoculation - slow healing ulcer, bacilli spread to lymphatics and bloodstream • 20% mortality if untreated • Respiratory anthrax is usually fatal even if treated • GI anthrax also occurs in some Asian countries • Excellent biological warfare agent eschar Gram stain with spores

8. Anthrax Pathogenesis • Poly-D-glutamic acid (protein) capsule – antiphagocytic • 3 component exotoxin • Protective antigen (PA) - binds to cells, forms channel that permits EF and LF to enter • Edema factor (EF) – adenyl cyclase causes fluid to accumulate at the site of infection and inhibit immune function • Lethal factor (LF) • disrupts cell's functions, stimulates TNF-alpha and IL-1-beta, kills infected cells

9. Anthrax Vaccine • Made from avirulent, nonencapsulated B. anthracis strain • Requires series of injections and annual boosters • Used in military

10. Listeria monocytogenes • Gram-positive bacilli/coccobacilli • Catalase-positive • Motile • Esculin positive •  hemolytic • Multiplies at 4 o C Umbrella motility

11. Habitat & Sources of Infection • Intestines of mammals, birds • Cheese, other dairy products • Vegetables • Undercooked meat • Epidemiology & Disease • Asymptomatic carriage • Meningitis/sepsis in immunocompromised host • Foodborne illness • Neonatal infections (transplacental)

12. Listeria: Pathogenesis • Organism adapted to grow at low temperatures • Multiplies in cytoplasm of macrophages and epithelial cells • Listeriolysin O (hemolysin) • Asymptomatic carriage reservoir

13. Erisipelothrix rhusopathiae • Gram-positive bacillus • Common in animals • Transmitted to humans by skin wound • Occupational hazard in butchers, farmers, veterinarians • Self-limited skin lesions with erythema & eruption • May spread to bloodstream

14. Aerobic Actinomycetes • Gram-positive, catalase-positive rods • Occur in soil and decaying vegetation + mycolic acid - mycolic acid Nocardia Streptomyces Rhodococcus Tropheryma Others Mycobacterium Corynebacterium

15. Corynebacterium • Gram-positive, curved pleomorphic rods • “Chinese Letters” • Aerobic/facultatively anaerobic • Grow readily on Sheep blood agar • Catalase & oxidase positive • Usually non-motile • Commensals on skin • Some species pathogenic

16. Diphtheria (C. diphtheriae) • Spreads by droplet • Phospholipase D increases vascular permeability & promotes spread • Phage-coded exotoxin (A&B subunits) acts on respiratory mucous membranes interfering with protein synthesis by inactivating EF-2 • Pseudomembrane of fibrin, bacteria, epithelial & phagocytic cells impairs breathing • Toxin spreads to heart, CNS, & adrenals • Selective media (cysteine-tellurite) • Identify in Public Health Labs • Protect by toxoid vaccination - DaPT • < 5 cases/yr in US

17. Other Pathogenic Corynebacteria • C. jeikeium (JK) • Opportunistic bloodstream infections in bone marrow transplant recipients • Multiple antibiotic resistance - except vancomycin and tetracycline • Commonly carried on skin of hospitalized persons • C. urealyticum • Occasional cause of UTI & stones (splits urea)

18. Nocardia • Epidemiology • Ubiquitous in environment • Inhalation • Cutameous inoculation • Pathogenesis • Cord factor – prevents phagolysosome fusion & intracellular killing in phagocytes • Catalase • Superoxide dismutase Filamentous gram-positive bacilli Partial Acidfast Smear

19. Nocardia Diseases • Opportunist (HIV, Malignancy) • Pneumonia • Lung Abscess • Brain Abscess/Meningitis • Cellulitis/ulceration • Mycetoma Suppurative granulomas Fibrosis Necrosis Sinus tracts

20. Nocardia 6 year-old boy with HIV with 2-month-history of fever & cough.

21. Nocardia Lab Diagnosis • Stained smears • Culture - slow growth ~ 7 days in 5% CO2 • Several species associated with human disease • Identify by PCR Nocardia colonies Middlebook 7H11 agar Aerial hyphae

22. Rhodococcus equi • Gram-positive weakly acidfast bacilli that sometimes revert to coccoid forms • Common in animals & environment • Opportunistic lung infections in AIDS & transplant patients • Facultative intracellular pathogen that survives in macrophages, leading to granuloma & abscess Slow-growing, mucoid salmon-colored colonies of R. equi on chocolate agar

23. Tropheryma whippeli • Actinomycete etiologic agent of Whipple’s Disease – a malabsorptioin syndrome affecting the small bowel • Organism cannot be cultured except in tissue culture • Diagnose by histopathology & PCR