Ken B, Waites, M.D. F(AAM)

Professor of Pathology

Director of Clinical Microbiology

Gram-positive Bacilli

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

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

Bacillus Endospores

• Thick walled structures formed by vegetative cells

• Resistant to radiation, chemicals, heat, desiccation  (dipicolinic acid)

• Steam autoclaving necessary for destruction

Bacillus: Clinical Significance

• Common environmental commensals

• Occasional opportunistic pathogens & culture contaminants

• B. cereus group

• B. anthracis

• Others

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

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

Gram stain with spores

eschar

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

Anthrax Vaccine

• Made from avirulent, nonencapsulated B. anthracis strain

• Requires series of injections and annual boosters

• Used in military

Listeria monocytogenes

•Gram-positive bacilli/coccobacilli•Catalase-positive•Motile•Esculin positive hemolytic•Multiplies at 4 o C Umbrella

motility

• 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)

Listeria: Pathogenesis

• Organism adapted to grow at low temperatures

• Multiplies in cytoplasm of macrophages and epithelial cells

• Listeriolysin O (hemolysin)

• Asymptomatic carriage reservoir

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

Aerobic Actinomycetes

• Gram-positive, catalase-positive rods

• Occur in soil and decaying vegetation

+ mycolic acid - mycolic acid

MycobacteriumCorynebacterium

NocardiaStreptomycesRhodococcusTropherymaOthers

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

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

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)

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

Nocardia Diseases• Opportunist (HIV, Malignancy)

– Pneumonia– Lung Abscess– Brain Abscess/Meningitis– Cellulitis/ulceration– Mycetoma

Suppurative granulomasFibrosisNecrosisSinus tracts

Nocardia

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

Nocardia Lab Diagnosis

• Stained smears• Culture - slow

growth ~ 7 days in 5% CO2

• Several species associated with human disease

• Identify by PCRNocardia colonies Middlebook 7H11 agarAerial hyphae

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

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