Microbial World

Prokaryotes:1. Bacteria2. Archae

Eukaryotes1. Algae2. Fungi3. Protozoa4. Parasites

Fungi

Features: Cell wall: Chitin Cell membrane:

Ergosterol Zymosterol

Aerobic/ facultatively anaerobic Mostly microscopic Moisture essential for growth

Types of Fungi Yeasts:

Single celled Reproduction:

Budding

Molds: Long filaments

(hyphae) Form mycelium Septate/ Non

septate

Dimorphic

Importance Biologic recycling of organic matter Preparation of foods:

Beer Cheese Bread Wine Mushrooms

Economic impact: Plant diseases Source of biologically active compounds:

•Hallucinogens•Adrenergic alkaloids•Vitamins•Mutagens

•Carcinogens•Antibiotics •Immunosuppressive agents•Potential anticancer substances

Groups1. Zygomycetes:

Bread moulds (Rhizopus)

Food spoilage organisms

Rhizomucor

2. Basidiomycetes: Common

mushrooms Cryptococcus Malassezia

3. Ascomycetes: Aspergillus Histoplasma Coccidioides Candida Pneumocystis Sporothrix Dermatophytes

4. Deuteromycetes (Fungi Imperfectii):

Medically & economically imp fungi

Penicillin producing

Types of Fungal Diseases

1. Fungal allergies2. Mycotoxicoses3. Mycoses

Fungal Allergies Strong hypersensitivity reactions against:

Fungal spores Fungal components

Do not require: Growth Viability

Depending upon the site of deposition of allergens:

Rhinitis Sinusitis Bronchial asthma Alveolitis Generalized pneumonitis

Mycotoxicoses

Mycetismus Mycotoxins:

Amatoxins Phallotoxins Aflatoxin Ochratoxin Sporidesmin Zearalenone Sterigmatocystin

Target organ: Liver

Mycoses

Actual growth of a fungus on a human or animal host

Establishment of mycoses depends upon:

Host defenses Size of innoculum Route of exposure Virulence of the fungus

Clinical Classification of Mycotic Infections

Superficial: Pityriasis versicolor Tinea nigra

Cutaneous: Candidiasis Dermatophytosis

Subcutaneous: Rhinosporidiosis Rhinoentomophthoromyco

sis

Systemic: Histoplasmosis Paracoccidioidomycosis Candidiasis Cryptococcosis Aspergillosis Mucormycosis

Specimen Collection, Handling & Transport

Sample Collection: Primary criterion for diagnosis of mycotic infections Transportation & processing done ASAP

Tissue from site of active disease- ideal Most common specimens:

Respiratory secretions Hair Skin Nail Tissue Blood Bone marrow CSF

Respiratory Specimen

Viscous material (Tracheal aspirate): Cotton Swab Specimen digested with trypsin & concentrated

Sputum: Deep cough early in the morning Nebulizer to induce sputum

Collected into a sterile screw top container

Media: Non selective Media with antibiotics

KOH preparation

Mucin Collection

Nasal decongestant spray Flush with 20ml N/S Forceful exhalation through nose Return collected in sterile pan

Skin: 70% isopropyl alcohol before sampling Scraped from outer edge of a surface

lesion

Blood: Transport medium required

Exudates/ Pus: Sterile sealed container

Diagnosis

Direct microscopic examination

Culture Serology

Direct Microscopic Examination

Wet preparations: KOH mount India Ink Calcofluor white

Histologic stains: Periodic- Acid Schiff (PAS ) stain Grocott- Gomori methenamine silver nitrate

(GMS) stain H&E stain Giemsa stain Masson- Fontana stain

Culture

Culture Media: Saboraud’s Dextrose Agar SDA with antibiotics Brain Heart Infusion (BHI) agar enriched

with blood & antibiotics

Incubation: Temp: 25-30°C (37°C for dimorphic fungi) Duration: 4-6 weeks

Candida Candida: Part of normal flora of skin, mucus

membranes & GIT Candidiasis: Most common systemic mycoses Pathogenic strains:

C. albicans C. tropicalis C. glabrata C. krusei

Clinical Classification of Candidiasis: Cutaneous & Mucosal candidiasis:

Thrush Stomatitis Esophagitis

Systemic Candidiasis Esophagitis

Chronic Mucocutaneous Candidiasis

Predisposing factors: Cutaneous & mucosal

candidiasis: Physiologic:

Pregnancy Old age Infancy

Traumatic Hematologic: AIDS DM Iatrogenic:

Antibiotics Steroids

Systemic Candidiasis: Immunosuppression Surgery Steroids Malignancies Cytoxic drugs

Morphology Dimorphism:

Yeast cells True hyphae Pseudohyphae Germ tubes

Microscopy: Spherical/ ellipsoidal budding

yeasts Size: 3-6 μm Cornmeal agar: Chlamydiospores

Culture: Species cannot be differenciated Within 24-48hrs Raised Cream coloured Opaque 1-2mm Hyphae penetrating the agar

medium

Aspergillus

Ubiquitous molds Numerous species Approx 20 cause human infection Pathogenic species:

A. fumigatus A. flavus: Nose & PNS A. niger: systemic disease in

immunocompromised

Clinical diseases: Otomycosis Fungal rhinosinusitis

Morphology Microscopy:

Conidiophores Expand into large vesicles at the

end Covered with phialides

Culture: Powdery Pigmented

A. fumigatus: Gray, green A. flavus: Yellow- green A. niger: Black

Aspergillus niger & flavus

Mucormycosis Phycomycosis,

zygomycosis Molds Class: Zygomycetes Order: Mucorales Fungi:

Ubiquitous Thermotolerant Saprophytes

At risk patients: Acidosis Leukemia Immunocompromise

Etiologic agents: Rhizopus oryzae R. rhizopodiformis Absidia corymbifera R. pusillus Rhizomucor spss. Mucor spss.

Clinical manifestations: Rhinocerebral

mucormycosis Thoracic mucormycosis Cutaneous infections

Morphology

Microscopy: Broad Sparsely septate hyphae (10μm) Twisted & ribbonlike Branching at rt. angles

Culture: Rapid growth Abundant, cottony aerial mycelia

Paracoccidioidomycosis C/A: Paracoccidioides brasiliensis Chronic granulomatous disease:

Mucous membranes Skin Respiratory system

Most cases from Brazil Invade mucous memb of mouth→ teeth fall

out White plaques in buccal mucosa Histologically: Captain’s wheel

Cryptococcosis C. neoformans Distinctive yeast Diseases:

Meningitis Pulmonary disease

Found in pigeon & chicken droppings Diagnosis:

India ink test Latex agglutination test for cryptococcal

antigen

India Ink staining of CSF

Polyenes Azoles

Imidazoles Triazoles

Echinocandins

Allylamines Flucytosine Griseofulvin

Polyenes Eg.:

Amphotericin B Nystatin

MOA: Bind to sterols of eukaryotic cell memb→

leakage of cell contents Amphotericin B:

Active against all fungi Leishmania Given parenterally Poor CSF penetration

ADRs: Fever Rigor Nephrotoxicity Hyperkalemia Headache

Azoles Inhibit cyt p450 14α-demethylase

→inhibit fungal cell wall synthesis Active against:

Candida Dermatophytes Aspergillus

Imidazole: Topical: Clotrimazole Systemic: Ketoconazole

Triazoles: Fluconazole: Inactive against invasive

moulds Itraconazole: Inactive against zygomycetes

Echinocandins Capsofungin Inhibit cell wall glucan synthesis

→cell wall lysis Active against:

Candida Aspergillus

Inactive against: Other moulds Cryptococcus

Allylamines

Terbinafine Reduce ergosterol synthesis Active against

dermatophytes Uses:

Skin dermatophyte infection Nail dermatophyte infection

Flucytoscine: Incorporates into fungal mRNA instead of uracil

→ disruption to protein & DNA synthesis Activity:

Cryptococcus Candida

Resistance: Common ADRs:

Bone marrow toxicity Hepatotoxicity

Griseofulvin: MOA unclear Use: Nail infections

Mycotic Diseases of the External Ear

Otomycosis Dermatophytosis Chromoblastomycosis Sporotrichosis

Otomycosis

Defn: Superficial, diffuse, fungal infection of the ear canal

Predisposing condition usually present

Aetiological agents: Aspergillus: (Tropical & Subtropical

regions) Niger Flavus Fumigatus

Candida: (Temperate regions) Albicans Parapsilosis Tropicalis

Penicillium Rhizopus Mixed

Epidemiology

Environment: Warm Humid

Children less commonly affected

Not contagious Predisposing factors:

Seborrhic dermatitis Psoriasis Prolonged use of:

Topical antibiotics Topical corticosteroids

Clinical Manifestations C/C:

Aural fullness Pruritis Discharge

Otoscopy: Debris Erythematous/ oedematous ear

canal A. niger: (Blotting paper)

Mat of fungus Black sporing heads

Chronic infection: Eczematoid change Lichenification

Diagnosis

Clinical Microscopic

examination Culture

Management

Removal of debris Cleaning Antifungal agents:

Local application Gauze packs Mercurochrome & boric

acid

Mycotic diseases of the nose and nasal passages

Entomophthoramycosis Rhinosporidiosis

Entomophthoramycosis

Definition:Chronic localised subcutaneous fungal

infection that originates from nasal mucosa and spreads painlessly to the adjacent subcutaneous tissue of the face

Rare Seen in healthy individuals Severe facial disfigurement C/A: Conidiobolus coronatus

Management

Oral antifungal drugs Treatment continued 1mnth after

lesions have disappeared Surgical resection:

Hastens spread of infection

Rhinosporidiosis

Definition: Uncommon granulomatous infection that affects

nasal mucosa, ocular conjunctiva & other mucosa

Etiology: Rhinosporidium seeberi Fungi: controversial 18S small subunit ribosomal DNA: Mesomycetozoa

In tissues: Thick walled sporangium like structures Endospores

Epidemiology

Geographical distribution: South India Sri Lanka East Africa Central & South America

Natural habitat: Stagnant pools of fresh

water

M>F Age:15-40yrs

Clinical Features

Nasal obstruction Rhinoscopy:

Pink/ Red/ Purple Papular/ Nodular Smooth surfaced Papillomatous/ Proliferative

Diagnosis: HPE:

Large sporangia filled with spores

Thick wall Operculum

Rhinosporidiosis

Management: Surgical excision Cauterization

Outcomes & Complications:

Recurrence

Mycotic Diseases of Paranasal Sinuses

Classification (Based on HPE & C/F):1. Invasive Sinusitis:

1. Active Invasive2. Chronic Invasive3. Chronic granulomatous invasive or

paranasal granuloma

2. Noninvasive Sinusitis3. Allergic Fungal Sinusitis

Invasive Fungal Sinusitis

Diagnosis: Evidence of

sinusitis: Radiographic Nasal endoscopy

Fungal hyphae: HPE

Etiological Agents

Acute fulminant: Rhizopus spss.

R. arrhizus Absidia spss. Rhizomucor spss. Aspergillus spss.

A. flavus A. fumigatus

Fusarium spss. S. apiospermum

Chronic invasive: Alternaria spss. Aspergillus spss. Bipolaris spss. Curvularia spss. Exserohilum spss.

Granulomatous invasive:

A. flavus

Epidemiology Worldwide Adults Immunocompromised children Risk factors:

Prolonged neutropenia Metabolic acidosis Hematological malignancies Haematopoetic stem cell transplant

recipients Diabetics Corticosteroid therapy Deferoxamine treatment HIV infection

Clinical Features Acute Invasive:

Immunocompromised Unilateral facial swelling Unilateral headache Nasal obstruction/ pain Serosanguinous nasal

discharge Necrotic black lesions on:

Hard palate Nasal turbinate

Periorbital/ perinasal swelling Destruction of facial tissue Ptosis Proptosis Ophthalmoplegia Loss of vision

Chronic invasive: Nasal obstruction Chronic sinusitis Thick nasal polyposis Thick purulent mucus Orbital apex syndrome Cavernous sinus thrombosis

Chronic granulomatous:

Nasal obstruction Unilateral facial discomfort Enlarging mass Proptosis

Diagnosis

CT Scan: Acute invasive:

Multiple sinuses Unilateral No air fluid level Thickening of sinus

lining Bone destruction

Chronic invasive: Hyperdense mass Sinus wall erosion

Chronic granulomatous: Opacification of

sinuses Erosion

MRI: Cavernous sinus Cerebral

Local biopsy: HPE Direct microscopy:

KOH mount Culture

Management

Control of underlying host disorders Removal of necrotic & infected

tissue Effective antifungal therapy

Noninvasive Fungal Sinusitis

Fungal ball: Dense mass of fungal

hyphae Aetiological agent:

Aspergillus fumigatus Other Aspergillus spss S. apiospermum Alternaria

Epidemiology: Older age group F>M

Clinical Features Asymptomatic Nasal obstruction Purulent nasal

discharge Cacosmia Facial pain Unilateral symptoms Unusual symptoms:

Fever Cough Proptosis Epistaxis Diplopia Nasal polyp

Diagnosis CT Scan:

Partial/ total opacification Flocculent calcification

Mucopurulent material: HPE:

Dense matted fungal hyphae separate from but adjacent to the mucosa of sinus

No allergic mucin No granulomatous reaction No fungal invasion

Management: Surgical removal No antifungal agents

Outcomes & Complications:

Recurrence: Rare Intracerebral bleed/ infarct Invasive fungal sinusitis

Allergic Fungal Sinusitis Noninvasive

Immunocompetent individuals Chronic rhinosinusitis Criteria for diagnosis:

Chronic rhinosinusitis (CT Scan) Allergic mucin

Clusters of eosinophils Eosinophillic byproducts

Noninvasive fungal elements Type I (IgE mediated)

hypersensitivity Nasal polyposis

Ponikau et al. (1999): 210 pts with chronic rhinosinusitis Fungus in nasal mucus: 202 pts

(96%) Surgical treatment: 101 Allergic mucin: 97 (96%) Fungal elements in HPE: 82 (81%)

Conclusion: AFS- Underdiagnosed disorder

Aetiology: Aspergillus Dematiaceous environmental

moulds: Alternaria Bipolaris Cladosporium Curvularia Drechslera

Epidemiology: Young immunocompetent

adults Relapsing rhinosinusitis Unresponsive to:

Antibiotics Antihistamines Corticosteroids

M=F Atopic Southern United States

Clinical Features h/o Chronic rhinosinusitis U/L nasal polyposis Thick yellow-green mucus Bone necrosis of thin walls of sinus Proptosis DNS to opposite side Pt with nasal polyposis responding

only to oral corticosteroids

Diagnosis CT Scan: Serpiginous opacification of >1 sinus Mucosal thickening Bone erosion No tissue invasion

Microscopic Examination of allergic mucin:

Eosinophils Fungal elements

Histologic examination to r/o invasion Lab tests:

Eosinophilia Total serum IgE Specific IgE against fungal Ags +ve skin prick tests

Fungal cultures

AFS

Management Surgical debridement Adjunctive medical

management: Oral corticosteroids Specific allergen immunotherapy Nasal corticosteroids Antihistamines Antileukotrienes Sinonasal saline lavage

Systemic antifungals: not effective

Mycotic Diseases of the Throat

Candidiasis Histoplasmosis Paracoccidioidomycosi

s Blastomycosis Coccidioidomycosis Cryptococcosis

Candidiasis

Infections caused by organisms of genus Candida

Etiological Agents: C. albicans C. glabrata C. krusei C. tropicalis C. parapsilosis

Epidemiology C. albicans:

Commensal in the mouth of 40% ppl No. ↑es with:

Tobacco smoking Dentures

Host factors: General:

Debilitated pts.: Broad spectrum antibiotics Corticosteroids DM Severe nutritional deficiencies Immunosuppressive diseases eg AIDS

Local: Trauma:

Unhygienic dentures Ill fitting dentures

Tobacco smoking

Clinical Manifestation

Clinical forms: Pseudomembranous Erythematous (or atrophic) Hyperplastic (or

hypertrophic)

Pseudomembranous Pts using steroid inhalers Immunocompromised individuals Neonates Terminally ill pts Lesions:

Raised white Surface of:

Tongue Soft & hard palate Buccal mucosa Tonsils

Confluent plaques Painless

Throat involvement: Severe dysphagia Pseudomembrane wiped off:

Pseudomembranous Candidiasis

Candidiasis

Erythematous Associated with:

Broad spectrum antibiotic treatment

Chronic corticosteroid use HIV

Any part of oral mucosa Lesions:

Flat Red Tongue: depappillated areas

Hyperplastic (Candida leukoplakia)

Lesions undergo malignant transformation

Lesions: Small, palpable, translucent white areas Large, dense, opaque plaques, hard,

rough

Lesions cannot be removed Site:

Inner surface of both cheeks Tongue

Other Candidal Lesions Chronic atrophic

candidiasis: Denture stomatitis Associated with oral prostheses Asymptomatic Soreness Cheilitis

Laryngeal Candidiasis: Hoarseness Dysphagia Stridor Plaques on laryngeal mucosa

Diagnosis: Clinical Microscopy HPE Culture

Management:

Antifungals Topical Systemic

Mycotic Colonization of Tracheo-oesophageal Voice

Prostheses Biofilm formation Invasion of silastic Causative agents:

C. albicans C. glabrata C. krusei C. tropicalis

Results in: Valve failure Device replacement

Local antifungal therapy: inadequate

Metal coating of prostheses

Thank you