respiratory mycoses (reduced version) dr elizabeth m. johnson, director hpa mycology reference...
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Respiratory mycosesRespiratory mycoses(reduced version)(reduced version)
Dr Elizabeth M. Johnson, DirectorDr Elizabeth M. Johnson, Director
HPA Mycology Reference Laboratory, HPA Mycology Reference Laboratory,
BristolBristol
Kingdom FungiKingdom Fungi
Eucaryotic organismsEucaryotic organisms No chlorophyllNo chlorophyll Absorb nourishment from living or dead organic Absorb nourishment from living or dead organic
materialmaterial Filamentous form: septate or aseptate hyphaeFilamentous form: septate or aseptate hyphae Yeasts: unicellular fungiYeasts: unicellular fungi Sexual and asexual formsSexual and asexual forms
Inhalation of fungal sporesInhalation of fungal spores
Spores Spores 8 µm8 µm
SINUSITISSINUSITIS
PULMONARYPULMONARYASPERGILLOSISASPERGILLOSIS
DISEASE SPECTRUMDISEASE SPECTRUM
ALLERGYALLERGY
ASPERGILLOMAASPERGILLOMA
DISEMINATEDDISEMINATEDASPERGILLOSISASPERGILLOSIS
2-4 µm2-4 µm
Schizophyllum communeSchizophyllum commune
Basidiomycete often found on rotting woodBasidiomycete often found on rotting wood
Increasingly recognised as a cause of Increasingly recognised as a cause of sinusitissinusitis
Produces raised fruiting bodies on prolonged Produces raised fruiting bodies on prolonged incubationincubation
Look for clamp connections and spiculesLook for clamp connections and spicules
Presence of risk factorsPresence of risk factors
Clinical signsClinical signs
LaboratoryLaboratory
Microscopy & Culture of sputumMicroscopy & Culture of sputum
and BALand BAL
Serology - detection of antibodies Serology - detection of antibodies and fungal antigens and fungal antigens
Detection of fungal DNA Detection of fungal DNA using PCR using PCR
Diagnosis of respiratory mycosesDiagnosis of respiratory mycoses
Patchy changes seen on chest radiograph of Patchy changes seen on chest radiograph of patient with invasive aspergillosispatient with invasive aspergillosis
Aspergillosis: radiological findingsAspergillosis: radiological findingsin the lungsin the lungs
Halo sign on CT scan - early finding (1-10 days)Halo sign on CT scan - early finding (1-10 days)
Crescent-shaped lesions on chest radiograph Crescent-shaped lesions on chest radiograph
- late sign following neutrophil recovery- late sign following neutrophil recovery
Halo sign Halo sign Air crescent Air crescent
Aspergillus Aspergillus hyphae in a hyphae in a sputum samplesputum sample
Microscopy of a Microscopy of a mucoraceous mouldmucoraceous mould
Absidia corymbiferaAbsidia corymbifera
Fungal serology - AntigensFungal serology - Antigens
Latex agglutination + Latex agglutination + ++++++
ELISA ELISA + + ++++++
Tests for antigenaemiaTests for antigenaemia Asp Cryp Asp Cryp
TARGET
• Cell wall componentsCell wall components::
mannans mannans
galactomannangalactomannan
1,3 beta-D-glucans1,3 beta-D-glucans
• Cytoplasmic antigens Cytoplasmic antigens • MetabolitesMetabolites
Aspergillus ELISAAspergillus ELISARat EBA-2 monoclonal antibodyRat EBA-2 monoclonal antibodydirected against galactomannandirected against galactomannan
sensitivity - 1 ng/mlsensitivity - 1 ng/ml
G - testG - testAmebocyte limulus lysate test Amebocyte limulus lysate test Horse shoe crab coagulation Horse shoe crab coagulation
cascadecascadesensitivity - 1 pg/mlsensitivity - 1 pg/ml
Pitfalls of PCRPitfalls of PCR
Choice of sampleChoice of sampleBlood, Serum or PlasmaBlood, Serum or PlasmaBALBALTissueTissue
Method of DNA extractionMethod of DNA extractionMechanicalMechanicalChemicalChemicalEnzymesEnzymes
Contamination:Contamination:Environmental Environmental AmpliconAmplicon
Absence of standardization of reagents and targetsAbsence of standardization of reagents and targets
Infection with Infection with Scedosporium Scedosporium spp.spp.
Pseudallesheriosis - sexual form (teleomorph) Pseudallesheriosis - sexual form (teleomorph) Pseudallescheria boydiiPseudallescheria boydii
Clinically indistinguishable from invasive Clinically indistinguishable from invasive aspergillosisaspergillosis
Specific features:Specific features:
Infection often follows near-drowning Infection often follows near-drowning accidentsaccidents
Neutropenia is a risk factorNeutropenia is a risk factor
Refractory to treatmentRefractory to treatment
Scedosporium apiospermumScedosporium apiospermum
Scedosporium prolificans
Infection with Infection with Fusarium Fusarium spp.spp.
Clinically very similar to invasive aspergillosisClinically very similar to invasive aspergillosis
Specific features:Specific features:
Cutaneous lesions (70%)Cutaneous lesions (70%)
Blood culture often positive (50-70%)Blood culture often positive (50-70%)
Often water borneOften water borne
Sinus infection may lead to disseminated diseaseSinus infection may lead to disseminated disease
Fusarium solaniFusarium solani
Cryptococcosis in the UK Cryptococcosis in the UK 1980 - 19991980 - 1999
0
10
20
30
40
50
60
1980 1981 1982 1983 1984 1985 1986 1987 1988 1989 1997 1998 1999
Non-AIDS AIDS
Year
No. cases
AIDS-defining 24 18 24 illness
Knight et al. 1993 J Infect 27:185-191
Cryptococcus neoformansCryptococcus neoformans
CAN infect healthy, non-immunocompromised hostsCAN infect healthy, non-immunocompromised hosts
Today mainly associated with HIV-infected patientsToday mainly associated with HIV-infected patients
AIDS-defining illnessAIDS-defining illness
HAART has had an impactHAART has had an impact
Mode of infection is inhalation but haematogenous Mode of infection is inhalation but haematogenous dissemination leads to meningitisdissemination leads to meningitis
Diagnosis by direct microscopy or LPA antigen testDiagnosis by direct microscopy or LPA antigen test
Indian ink stain of Indian ink stain of Cryptococcus neoformansCryptococcus neoformans
Pneumocystis carinii Pneumocystis carinii var. var. hominishominis= Pneumocystis jiroveci= Pneumocystis jiroveci
Coccidioides immitisCoccidioides immitis
Cause of coccidioidomycosisCause of coccidioidomycosis
Endemic areas southwestern USA, Endemic areas southwestern USA,
Central and South AmericaCentral and South America
Infection by inhalation of arthroconidiaInfection by inhalation of arthroconidia
Mild transient pulmonary infectionMild transient pulmonary infection
1% develop disseminated disease 1% develop disseminated disease
Coccidioides Coccidioides immitis immitis distributiondistribution
Endemic areas: Endemic areas: southwestern USA southwestern USA Central and South Central and South AmericaAmerica
From: From: Medical MycologyMedical MycologyKwon-Chung and BennettKwon-Chung and Bennett1992 Lea & Febiger1992 Lea & Febiger
Coccidioides immitis life cycleCoccidioides immitis life cycle
From: Medical Mycology Kwon-Chung and Bennett 1992 Lea & FebigerFrom: Medical Mycology Kwon-Chung and Bennett 1992 Lea & Febiger
Direct microscopyDirect microscopy
Endosporulating spherulesEndosporulating spherules
seen on direct microscopyseen on direct microscopy
of pus, sputum or joint of pus, sputum or joint
fluidfluid
Difficult to obtain in cultureDifficult to obtain in culture
CultureCulture
Mycelial colonies after Mycelial colonies after
incubation at 25 - 30ºC incubation at 25 - 30ºC
for 2-7 daysfor 2-7 days
Growth on actidioneGrowth on actidione
Typical arthroconidiaTypical arthroconidia
Exoantigen test to confirmExoantigen test to confirm
Gene-probe also availableGene-probe also available
Laboratory diagnosis of Laboratory diagnosis of coccidioidomycosiscoccidioidomycosis
Serology Serology Immunodiffusion and complement fixationImmunodiffusion and complement fixation
HistoplasmosisHistoplasmosis
Guano-enriched soil sourceGuano-enriched soil source
Tissue form: budding yeastsTissue form: budding yeasts
Reticuloendothelial systemReticuloendothelial system
Chronic cases - calcifications Chronic cases - calcifications
Widespread geographicallyWidespread geographically
- especially Eastern USA, & Africa- especially Eastern USA, & Africa
HistoplasmaHistoplasma var. var. capsulatum capsulatum and var. and var. duboisiiduboisii
HistoplasmosisHistoplasmosis
Direct microscopyDirect microscopy
Small, oval, budding yeast Small, oval, budding yeast
cells seen in Giemsa cells seen in Giemsa
stained smearsstained smears
Cells often seen withinCells often seen within
macrophagesmacrophages
CultureCulture
Mycelial colonies after Mycelial colonies after
incubation at 25 - 30ºC incubation at 25 - 30ºC
for 2-7 daysfor 2-7 days
Conversion to yeast on Conversion to yeast on
blood agar or BHI agar at blood agar or BHI agar at
37ºC, 2-6 weeks37ºC, 2-6 weeks
Laboratory diagnosis of histoplasmosisLaboratory diagnosis of histoplasmosis
Serology Serology
Immunodiffusion and Immunodiffusion and complement fixationcomplement fixation
Sensitive and specificSensitive and specific High positive predictive valueHigh positive predictive value High negative predictive valueHigh negative predictive value Simple, rapid and inexpensiveSimple, rapid and inexpensive
The ideal diagnostic testThe ideal diagnostic test