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MYCOBACTERIAMYCOBACTERIACORYNEBACTERIACORYNEBACTERIA

LEGIONELLALEGIONELLA

Faculty: Dr. Alvin Fox

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KEYWORDS• Acid FastAcid Fast• Tuberculosis (TB)Tuberculosis (TB)• M. tuberculosis M. tuberculosis (MDR, XDR)(MDR, XDR)• M. avium - M. intracellulareM. avium - M. intracellulare complex complex• M. bovis M. bovis • M. lepraeM. leprae• TubercleTubercle• PPDPPD• TuberculinTuberculin• MycobactinMycobactin• Cord factorCord factor• BCGBCG• Leprosy (Hansen's Disease)Leprosy (Hansen's Disease)• AIDS and TBAIDS and TB• Runyon groupsRunyon groups• Mycolic acidsMycolic acids

• DiphtheriaDiphtheria• C. diphtheriaeC. diphtheriae• Loeffler's agarLoeffler's agar• Tellurite agarTellurite agar• Metachromatic bodiesMetachromatic bodies• Diphtheria toxinDiphtheria toxin• Schick testSchick test• DiphtheroidsDiphtheroids• Legionella pneumophilaLegionella pneumophila• Legionnaires diseaseLegionnaires disease• Pontiac feverPontiac fever• Legionella micdadeiLegionella micdadei• Pittsburg pneumoniaPittsburg pneumonia

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Mycobacterium tuberculosisMycobacterium tuberculosis

obligate aerobeobligate aerobeacid-fast rodsacid-fast rods

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• M. tuberculosisM. tuberculosis• major human diseasemajor human disease

– healthy peoplehealthy people • problems problems

– association with AIDSassociation with AIDS– multiple drug-resistancemultiple drug-resistance

Tuberculosis Tuberculosis (TB, consumption)(TB, consumption)

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M. avium- M. intracellulareM. avium- M. intracellulare complex (complex (M. aviumM. avium))

• non-AIDSnon-AIDS– infection almost never infection almost never

• AIDS AIDS – major bacterial opportunistmajor bacterial opportunist

• multiple drug-resistance multiple drug-resistance

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• spread from cattlespread from cattle

• infected cattle are culledinfected cattle are culled– positive skin testpositive skin test

• rarely seen in USrarely seen in US

M. bovisM. bovis

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M. leprae

• leprosyleprosy

• major disease of third worldmajor disease of third world

• rare in US rare in US

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Transmission -tuberculosisTransmission -tuberculosis

• M. tuberculosis M. tuberculosis causes diseasecauses disease–healthy individuals healthy individuals

• transmitted man-man transmitted man-man

• airborne dropletsairborne droplets

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Pathogenesis of tuberculosisPathogenesis of tuberculosis

• infects lunginfects lung

• distributed within macrophagesdistributed within macrophages

• facultative intracellular pathogenfacultative intracellular pathogen– inhibits phagosome-lysosome fusioninhibits phagosome-lysosome fusion

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Cell-mediated immunity -Cell-mediated immunity -tuberculosistuberculosis

• infiltration infiltration – macrophages macrophages

– lymphocyteslymphocytes

• granulomasgranulomas

• tuberculestubercules

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Laboratory diagnosis - Laboratory diagnosis - tuberculosistuberculosis

• skin testingskin testing– delayed hypersensitivity delayed hypersensitivity

– tuberculin tuberculin

– protein purified derivative, PPDprotein purified derivative, PPD

• X-rayX-ray

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Positive skin test -tuberculosisPositive skin test -tuberculosis

• indicates exposure to organismindicates exposure to organism

• does not indicate active diseasedoes not indicate active disease

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Other minor pathogenesis factors Other minor pathogenesis factors tuberculosistuberculosis

• mycobactin mycobactin – siderophoresiderophore

• cord factor cord factor – damages mitochondriadamages mitochondria

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Laboratory diagnosis Laboratory diagnosis M. tuberculosisM. tuberculosis

• acid fast bacteria acid fast bacteria – sputum sputum

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Laboratory diagnosis Laboratory diagnosis M. tuberculosis M. tuberculosis (culture)(culture)

• grows very slowlygrows very slowly– two weeks or longertwo weeks or longer– non-pigmented coloniesnon-pigmented colonies– niacin productionniacin production

*differentiates from other mycobacteriadifferentiates from other mycobacteria

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TuberculosisTuberculosis

• polymerase chain amplificationpolymerase chain amplification– rapid diagnosisrapid diagnosis

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Antibotic treatment - Antibotic treatment - tuberculosistuberculosis

• extensive time periods (e.g. 9 months)extensive time periods (e.g. 9 months)

• organism grows slowly, or dormantorganism grows slowly, or dormant

• two or more antibioticstwo or more antibiotics

– e.g. rifampin and isoniazide.g. rifampin and isoniazid– resistance minimizedresistance minimized

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Tuberculosis and Drug resistance

• Multiple drug resistant (MDR)

– resistant to first line drugs

• Extremely drug resistant (XDR)

– Resistant to some of the second line drugs

– Nearly un-treatable

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VaccinationVaccination

• BCG vaccine BCG vaccine – an attenuated strain of an attenuated strain of M. bovisM. bovis – not effectivenot effective

• in US, in US, – incidence is lowincidence is low– vaccination not practicedvaccination not practiced– immunization interferes with diagnosisimmunization interferes with diagnosis

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Mycobacterium lepraeMycobacterium leprae

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Leprosy (Hansen's Disease)Leprosy (Hansen's Disease)• M. lepraeM. leprae

– causative agentcausative agent

• chronic disease chronic disease – disfigurementdisfigurement

• rarely seen in the U.S. rarely seen in the U.S.

• common in third worldcommon in third world• millions of cases millions of cases

• infects the skin infects the skin – low temperaturelow temperature

22ulcers, resorption of bone ulcers, resorption of bone worsened from careless use of hands (nerve damage)worsened from careless use of hands (nerve damage)

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• tuberculoidtuberculoid• few organisms few organisms • active cell-mediated immunityactive cell-mediated immunity

• lepromatous lepromatous • many organisms many organisms • immunosuppressionimmunosuppression

LeprosyLeprosy

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• in vitroin vitro – unculturable unculturable

• in vivoin vivo growth growth– low temperaturelow temperature– armadillo armadillo – mouse footpadmouse footpad

Production of Production of M. leprae M. leprae antigens and antigens and pathogenesis studiespathogenesis studies

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• lepromin lepromin – skin testingskin testing

• acid-fast stains acid-fast stains – skin biopsiesskin biopsies

• clinical picture clinical picture

LeprosyLeprosy

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Other mycobacterial species Other mycobacterial species (including (including M. aviumM. avium))

• infect immunocompromised hostinfect immunocompromised host• not transmitted man-man, healthy peoplenot transmitted man-man, healthy people• M. aviumM. avium – common – common• Other species - rareOther species - rare

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• tuberculosis-like tuberculosis-like • leprosy-likeleprosy-like

Mycobacterial diseasesMycobacterial diseases

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• M. avium M. avium is much less virulent than is much less virulent than M. tuberculosisM. tuberculosis– does not infect healthy peopledoes not infect healthy people– infects AIDS patientsinfects AIDS patients

• M. aviumM. avium infects infects– when CD4 (helper T cell) count greatly decreasedwhen CD4 (helper T cell) count greatly decreased

• M. tuberculosisM. tuberculosis infection infection– infects healthy peopleinfects healthy people– infects AIDS patientsinfects AIDS patients

* earlier stage of diseaseearlier stage of disease* more systemicmore systemic

Mycobacteria and AIDSMycobacteria and AIDS

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• systemic disease (versus pulmonary)systemic disease (versus pulmonary)– greater in AIDS greater in AIDS

• lesions often lepromatouslesions often lepromatous

Clinical features with AIDSClinical features with AIDS

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• selected primarily for selected primarily for M. tuberculosisM. tuberculosis

• if if M. aviumM. avium involved other antibiotics included involved other antibiotics included

Antibiotic therapy Antibiotic therapy

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Other speciesOther species

• pigmented or notpigmented or not• pigmentation pigmentation

– in the light in the light – in the darkin the dark

• growthgrowth– fastfast– slow slow

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• cellular fatty acid profilescellular fatty acid profiles• mycolic acid profilesmycolic acid profiles• genetic markersgenetic markers

Mycobacterial species identificationMycobacterial species identification

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Mycolic acidsMycolic acids• mycobacteria mycobacteria – longest chain lengthlongest chain length– strongly acid faststrongly acid fast

• nocardianocardia – intermediate chain lengthintermediate chain length– weakly acid fastweakly acid fast

• corynebacteria corynebacteria • shortest chain lengthshortest chain length– not acid fastnot acid fast

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Corynebacterium diphtheriaeCorynebacterium diphtheriae

Gram positiveGram positivestrict aerobestrict aerobepleomorphic (e.g. club-shaped)pleomorphic (e.g. club-shaped)

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• infection infection – upper respiratory tract (pharynx)upper respiratory tract (pharynx)– pseudomembrane pseudomembrane – chocking chocking – bacteria do not spread systemicallybacteria do not spread systemically

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DiphtheriaDiphtheria

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This child has diphtheria resulting in a thick gray coating over back of throat. This coating can eventually expand down through airway and, if not treated, the child could die from suffocation CDC

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Diptheria toxinDiptheria toxin

• spreadsspreads

• systemic and fatal injury systemic and fatal injury

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• B binds to host cellB binds to host cell• A inhibits protein synthesisA inhibits protein synthesis• ADP-ribose moiety (NADH) ADP-ribose moiety (NADH) attaches attaches

– elongation factor 2 inhibited elongation factor 2 inhibited

Diphtheria toxinDiphtheria toxin

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TreatmentTreatment

• anti-toxinanti-toxin• antibioticantibiotic

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• diseasedisease vanished in US vanished in US – without immunization will returnwithout immunization will return

• toxoid (+ pertussis and tetanus) DPT toxoid (+ pertussis and tetanus) DPT • neutralizing antibodiesneutralizing antibodies

• colonization not inhibited colonization not inhibited – found in normal florafound in normal flora

Immunization against diphtheria Immunization against diphtheria (infant)(infant)

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• Schick skin testSchick skin test-toxin

Testing immunity

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Diphtheria toxinDiphtheria toxin

• coded by bacteriophage tox genecoded by bacteriophage tox gene– not synthesized if iron presentnot synthesized if iron present

– iron-repressor complex forms iron-repressor complex forms

– inhibits expression of tox geneinhibits expression of tox gene

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• growth Loeffler's medium growth Loeffler's medium • stain for polyphosphate granulesstain for polyphosphate granules• metachromaticmetachromatic

– polyphosphate granules (pink) cell (blue) polyphosphate granules (pink) cell (blue)

• tellurite agartellurite agar• reduction by bacteria reduction by bacteria • tellurium precipitationtellurium precipitation• black coloniesblack colonies

Identification - Identification - C. diphtheriaeC. diphtheriae

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Identification – Identification – Exotoxin productionExotoxin production

• in vivoin vivo

• in vitroin vitro

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• diphtheroidsdiphtheroids– other corynebacteriaother corynebacteria– propionibacteriapropionibacteria

C. diphtheriae C. diphtheriae should not be confusedshould not be confused with: with:

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Legionella pneumophila

•Gram negative rodGram negative rod•Stains poorlyStains poorly

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• Legionnaire's diseaseLegionnaire's disease– pneumoniapneumonia– fatal if not treatedfatal if not treated– erythromycin erythromycin

• Pontiac feverPontiac fever– flu-likeflu-like– milder milder – self-limiting self-limiting – myalgia (muscle pain) myalgia (muscle pain) – no pneumonia no pneumonia

Legionella pneumophilaLegionella pneumophila

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Transmission

• contaminated air – infected water supply

• not spread person-person

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• environmental reservoir environmental reservoir – pools of stagnant waterpools of stagnant water– hot water towers hot water towers – air conditioning systemsair conditioning systems– intracellular growth, protozoaintracellular growth, protozoa

• facultative intracellular pathogenfacultative intracellular pathogen– humanshumans

Legionella pneumophiaLegionella pneumophia

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• small epidemics small epidemics – healthy peoplehealthy people

• more frequentlymore frequently– sick or elderlysick or elderly– hospitalshospitals– immunocompromisedimmunocompromised

Legionnaires diseaseLegionnaires disease

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• no growth conventional agarno growth conventional agar– e.g. sheep blood agare.g. sheep blood agar

• media must contain media must contain – iron iron – cysteine cysteine – e.g. charcoal yeast extract agare.g. charcoal yeast extract agar

• isolation clinical specimensisolation clinical specimens– poor poor

Identifcation of LegionellaIdentifcation of Legionella

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• Legionella pneumophilaLegionella pneumophila – several serogroupsseveral serogroups– most infections, serogroup 1most infections, serogroup 1

• Legionella micdadeiLegionella micdadei– Pittsburg pneumoniaPittsburg pneumonia

• OtherOther Legionella Legionella – occasional diseaseoccasional disease– legionellosislegionellosis

LegionellaeLegionellae

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• fatty acid profilingfatty acid profiling• DNA-DNA hybridizationDNA-DNA hybridization• 16S rRNA sequence16S rRNA sequence

• antigen detection in urineantigen detection in urine• serologyserology

Identification of Identification of LegionellaLegionella