UNP-MEDICINE 2021 microbiology

MYCOLOGY
LECTURER: DR. ZARA

May 1, 2019

HEAD NOTES  Highest incidence: candidiasis, dermatophytosis

 TITLE/EMPHASIZED H. capsulatum- fungal organism endemic in Mississippi and Ohio, USA.
 AUDIO
 NOTES/RECALLS

FUNGI
 Eukaryotic; most are nonmotile and possess a rigid wall -glucan
 Non photosynthetic
 <400 species are medically important
 <50 species cause >90% of the fungal infections of humans and
other animals
 Reside in nature and essential in breaking down and recycling
organic matter
 Contribute to the production of food and spirits (cheese, bread, beer)
 Provide useful bioactive secondary metabolites (antibiotics-
penicillin;
 immunosuppressive drugs – cyclosporine)
 Phytopathogens
Fungi- common among Asians who are immunocompromised; healthy
individuals will seldom have fungi; wet loving; likes to thrive best in wet
environment ex. repeated wearing of used socks for couple of days could
entertain fungal infection of foot (tinea pedis). Other fungal infections:
tinea unguium- nail biting, yellow discoloration of nails.
Tinea corporis- in the back of our body.
Fungi are eukaryotic (with organelles, mitochondria and ribosomes);
fungi possess very rigid cell wall glucan. Unlike in bacterias like
chlamydia and Mycoplasma which do not have cell wall.
Fungi are nonphotosynthetic and heterotrophic (feeding on dead organic
matter).
Alexander Fleming- discovered Penicillin. He placed penicillin in growth
of Staphylococcus and the bacteria was inhibited.
Cyclosporine- calcineurin inhibitor.
Table from Jawetz:

Ptyriasis versicolor- cause of your an-an; spaghetti or meatball

appearance in biopsy.
White Piedra- white portion of hair
Black Piedra- darkening/ blackening portion of hair. Superficial mycoses
are uncommon here in the Philippines.
Dermatophytosis- these are the fungal infections.
C. albicans- much common; opportunistic.
Candida infection of nail- Tinea unguium, groin- Jock itch, scalp- tinea
capitis, body- Tinea corporis
Endemic (Primary/systemic) mycoses can affect entire body
Sporothrix schenkii- “rose gardener’s disease”; infection from thorns of
roses.
Mycetoma- elephantiasis-like infection; most commonly caused by
Madurella mycetomatis; affects lower extremities.
*REMEMBER! Ptyriasis versicolor, Candidiasis, Sporothrichosis
and Mycetoma.
 Each fungal cell has at least one nucleus with a nuclear membrane, Systemic Candidiasis are very important. Candida can get into the blood
ER, (candidemia). There are 4 species of Candida. The most important is
 mitochondria and secretory apparatus Candida albicans.
 Most are obligate or facultative aerobes Cryptococcus neoformans- yeast; acquired from pigeon droppings; tested
 Chemotrophic ex. carbon sources and nitrogen using India Ink; the droppings when inhaled can disseminate into the
lungs and in the brain causing fever, headache and other meningeal signs.
MYCOSES Personal and social history are very important in fungal infection.
 Fungal infections Cryptococcus gattii- fungus from tree; uncommon.
 Most pathogenic fungi are exogenous -coming from the environment

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Aspergillus- common in molds; Aspergillus flavus produces aflatoxin and
associated with eating contaminated beans.
*Important opportunistic mycoses: Systemic candidiasis,
Cryptococcus and Aspergillus which are also common here in the
Philippines.
Mucormycosis- common
Pneumocystis jirovecii- formerly known as Pneumocystis carinii. In
Pneumocystis carinii infection there is increased LDH but nonspecific.
Check for whitish plaque in tongue, shortness of breath and CD4 count is
<200.
MICROBIOLOGY
 Most reproduce by budding
 Some produce buds that characteristically fail to detach and become
elongated (pseudohyphae – chain of elongated yeast cells)
 Colonies usually soft, opaque, 1-3 mm in size, cream-colored
 Yeast species are identified on the basis of physiologic tests, and
a few key morphologic differences
C. neoformans yeast cells (blue image)

GENERAL PROPERTIES AND CLASSIFICATION OF FUNGI

 Fungi grow in two basic forms:

1. YEASTS- grow as single cells that reproduce by asexual budding
2. MOLDS- production of multicellular filamentous colonies
branching cylindric tubules (hyphae) – 2-10 mm
mycelium- mass of intertwined hyphae that accumulates during
active growth
some hyphae are divided into cells by cross-walls (septa),
typically form at regular intervals during hyphal growth

 Vegetative or substrate hyphae – penetrate the surrounding

medium, anchor the colony and absorb the nutrients
 Aerial hyphae – project above the surface of the mycelium; usually
bear the reproductive structures of the mold
 When a mold is isolated from a clinical specimen, its growth rate,
macroscopic appearance and microscopic morphology are usually
sufficient to determine its genus and species
 Most helpful phenotypic (physical characteristics of mold) features:
ontogeny and morphology of the asexual reproductive
Broad based yeast- ex. Basidiomycetes
spores or conidia
 Thermally dimorphic (form different structures at different
temperatures)
HYPHAE
 Essential rigid cell wall (determines shape and protects from
• Coenocytic /aseptate vs.
osmotic and environmental stress)
Septate
o Composed largely of carbohydrate layers (long chains of
• Hyaline vs.
polysaccharides), glycoproteins and lipids
dematiaceous
o Sugar polymers: Chitin (b-1,4-linked N- acetylglucosamine),
• Fine vs. broad
glucans (b-1,3-glucan, b-1,6-glucan, mannans (polymers of
Hyaline- non pigmented
mannose- a-1,6-mannose)
Dematiaceous- pigmented
 Essential rigid cell wall
 During infection, fungal cell walls exert important pathobiologic
properties-
CONIDIA– asexual reproductive spores (MITOSPORES) produced either
o surface component mediate attachment to host cells
from the transformation of a vegetative yeast or hyphal cell or from a
o bind to pattern recognition receptors on host cell
specialized conidiogenous cell, which may be simple or complex and
membranes
elaborate
o cell wall glucans and other polysaccharides activate
 May be formed on specialized hyphae (conidiophores). Hyphae
the complement cascade and provoke an
usually transforms into conidia
inflammatory reaction
 MICROCONIDIA – small
o release immunodominant antigens that may elicit
 MACROCONIDIA – large or multicellular
cellular immune responses and diagnostic antibodies
dematiaceous – brown or black pigment to the fungal colony (melanized
cell walls) pigemnted
 Most fungi are obligate aerobes, some are facultative anaerobes,
but none are obligate anaerobes
 All fungi require a preformed organic source of carbon
 Natural habitat of most fungi: environment, except Candida albicans,
which is part of the normal human flora

YEASTS– single cells, usually spherical to ellipsoid in shape, 3-15 mm

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CLASSIFICATION ACCORDING TO COLONY TYPE AT

DIFFERENT TEMPERATURES
Saborauds medium- used for fungal; contains antibiotic that inhibit other
 MONOMORPHIC OR MONOPHASIC – produce one type of gram (-) organisms.
colony at both room and incubation temperature
 Ex.  DIMORPHIC OR DIPHASIC – produce a yeast-like colony at 37
 Cryptococcus neoformans – yeast only °C or incubation temperature (YEAST PHASE) and a filamentous type
 Aspergillus species – filamentous only of colony at room temperature (MYCELIAL PHASE) Ex:
 Candida albicans – yeastlike colony  Sporothrix schenckii
 Histoplasma capsulatum
 Blastomyces dermatitidis

ex. of filamentous or mold colony- Aspergillus

Histoplasma capsulatum- seen in bat droppings (in caves); the

manifestation of histoplasma is the same with manifestations of patients
with Pulmonary tuberculosis. One of your differentials here is Pulmonary
TB. If the patient went to endemic area, highly consider this organism.

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4. sporangiospores
TAXONOMY
 PHYLUM GLOMERULOMYCOTA, ORDER MUCORALES
 PHYLUM ASCOMYCOTA (Ascomycetes)
 65% of the known fungi, 85% of the human pathogens
 PHYLUM BASIDIOMYCOTA (Basidiomycetes)
 PHYLUM GLOMERULOMYCOTA, ORDER MUCORALES
 Sexual reproduction results in a ZYGOSPORE; asexual
reproduction occurs via SPORANGIA
 Vegetative hyphae sparsely septate
 Rhizopus, Lichtheimia, Mucor, Cunninghamella

PHYLUM ASCOMYCOTA (Ascomycetes)

 Sexual reproduction involves a sac (ascus) in which
karyogamy and meiosis occurs  ASCOSPORES; asexual
reproduction occurs via CONIDIA
 (+) septate hyphae
 Yeasts (Saccharomyces, Candida); molds (Coccidioides,
Blastomyces, Trichophyton)
PHYLUM BASIDIOMYCOTA (Basidiomycetes)
 Sexual reproduction results in dikaryotic hyphae and 4 progeny
BASIDIOSPORES supported by a club-shaped basidium; no
asexual spores
 Hyphae with complex septa; Mushrooms, Cryptococcus

DIAGNOSIS
 Species of a clinical isolate can be identified by molecular or
phenotypic methods (signature DNA sequences, morphology of
reproductive structures, physiologic properties)

SEXUAL SPORES
1. zygospores – single large spores with thick walls
2. ascospores – formed in a sac (ascus)
3. basidiospores – formed externally on a tip of a pedestal
(basidium)
FUNGI IMPERFECTI - Fungi that do not form sexual spores are “imperfect”

ASEXUAL SPORES
 arise by the differentiation of spore bearing hyphae without
nuclear fission
 most fungi of medical interest propagate asexually by forming
asexual spores (conidia) from the sides or ends of
specialized structures
1. arthrospores
2. chlamydospores
3. blastospores

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 TERBINAFINE – allylamine drug; blocks ergosterol synthesis by
inhibiting squalene epoxidase

CONIDIA ACCORDING TO ARRANGEMENT

 SESSILE AND LATERAL - develops directly on the side of the
hypha with no conidiophore or stem
 EN GRAPPE – clustered
 PEDUNCULATE - develop from the end of a short conidiophore

--END—

MYCOLOGY
 Mycology is the Study of Fungi (Monera, Protoctista, Fungi,
Plantae, Animalia).
 Fungi are eukaryotic cells and as such contain nuclei,
mitochondria, ER, golgi, 80S ribosomes, etc., bound by a
plasma membrane.
 Note that fungal cell membranes contain ergosterol rather than
cholesterol.
 In addition, fungi possess a rigid cell wall containing chitin,
 glucans and other sugar polymers.

Fungi are classified as

•Yeasts - round/oval cells that divide by budding

GROWTH AND ISOLATION OF FUNGI •Moulds - tubular structures (hyphae) that grow by longitudinal
 Saboraud’s agar – traditional mycological medium extension and branching. A mass of hyphae is called a mycelium
 Contains glucose and modified peptone (pH 7.0
 Does not readily support the growth of bacteria
 Inhibitory mold agar
MYCOTOXINS
 Poisonous substances produced by fungi
 Cause acute or chronic intoxication and damage
 Produced by mushrooms (Amanita sp.) mycetismus
 Aflatoxin – Aspergillus flavus

ANTIFUNGAL THERAPY
 POLYENES – Amphotericin B, Nystatin
 Bind to ergosterol in the cell membrane
 FLUCYTOSINE – pyrimidine analog Diseases Caused by Fungi
 AZOLES – interfere with the synthesis of ergosterol  Fungal infections in normal healthy adults are confined to
 ECHINOCANDINS – inhibit the synthesis of cell wall b-glucan conditions such as mucosal candidiasis (e.g., thrush) and
 GRISEOFULVIN – interferes with microtubule assembly dermatophyte (tinea) skin infections (e.g., athlete's foot).

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 However, in the immunocompromised host, a variety of
normally mild or nonpathogenic fungi can cause potentially
fatal infections.
Fungal infections are classified depending on the degree of tissue
involvement and mode of entry:
1. Superficial - localized to the skin, hair and nails.
2. Subcutaneous - infection confined to the dermis, subcutaneous tissue,
or adjacent structures.
3. Systemic - deep infections of the internal organs.
4. Opportunistic - cause infection only in the immunocompromised.

1. Superficial Mycoses
The Dermatophytes
 The dermatophytes are not a specific fungus, but rather a 2. Subcutaneous Mycoses
short-hand label for three genera of fungi that commonly cause  Subcutaneous infections confined to the dermis, subcutaneous
skin disease (tinea). tissue, or adjacent structures; there is no systemic spread.
•Epidermophyton spp.  They tend to be slow in onset and chronic in duration.
•Trichophyton spp.  These mycoses are rare in the US and are primarily confined to
•Microsporum spp. tropical regions (the Americas, South Africa, Australia).
•tinea capitis  The ease of travel provides the means for unusual fungal
•tinea barbae infections to be imported into this country.
•tinea pedis
•tinea cruis

Lobomycosis
3. Systemic Mycoses
Tinea pedis
 Systemic mycoses are invasive infections of the internal organs.
“athletes foot”
Epidermophyton spp.  The organism typically gains entry via the lungs, GI tract, or
through intravenous lines.
Examples include:
•Histoplasmosis
•Coccidiomycosis
•Blastomycosis

Histoplasmosis
 caused by Histoplasma, a dimorphic fungus
(grows as a mould at 25°C and as a yeast form at 37°C)

Tinea capitis
Microsporum spp.  Histoplasma is endemic in the Ohio-Mississippi river basins,
where it is found in soil contaminated with bird droppings and
Ringworm, dermatophyte infection (zoophilic) bat excrement.
 The infection is acquired through inhalation of the mould form
and the lungs are thus the most frequently affected site.
 Chronic pulmonary infection is frequently associated with
preexisting chronic lung diseases (i.e.- emphysema).
 All stages of this disease may mimic tuberculosis.
 The majority of acute cases (50%-90%) follow a subclinical
course (asymptomatic to flu-like Sx).
 The spectrum of the disease is wide, however, varying from an
acute benign pulmonary infection to a chronic pulmonary
infection and even a fatal disseminated disease.

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 Dissemination and a fatal course are more common in
immunocompromised, children less than 2 years, the elderly.

The lesion on the nose resulted from dissemination from the lungs

Discoloration of the skin caused by Histoplasma capsulatum  About 25% of patients with disseminated disease have
menningitis.

Blastomycosis
 A disease caused by the dimorphic fungus Blastomyces
dermatitidis
 It is endemic in the southeastern and south United States.
 Infection is acquired via inhalation.
 At least 50% of primary infections are asymptomatic.
 An acute pulmonary disease indistinguishable from a bacterial
pneumonia may occur after 30-45 days post exposure.

Oral lesions following hematogenous dissemination

Coccidiomycosis
 An infection caused by the dimorphic fungus Coccidioides
immitis.
 The disease is endemic only in regions of the Western Skin lesion following dissemination from the lungs
Hemisphere (Arizona, California, New Mexico and Texas).
 Coccidioidomycosis is acquired from inhalation and an acute 4. Systemic Mycoses, Opportunistic
respiratory infection occurs 7 to 21 days.  Opportunistic fungi are normally of marginal pathogenicity, but
 Most patients (50%) are asymptomatic. can infect the immunocompromised host.
 Symptoms, when they occur, typically resolve rapidly.  Patients usually have some serious immune or metabolic
 Occasionally, infection may result in a chronic pulmonary defect, or have undergone surgery.
condition and/or disseminate to the menninges, bones, Examples include:
joints,subcutaneous, or cutaneous tissues. Aspergillosis
Candidosis
Cryptococcosis

CANDIDIASIS

- an infection caused by a Candida spp.

- Antibiotic treatment can alter the normal bacterial flora
allowing Candida to flourish.
- Systemic candidiasis is common in the immunocompromised
(AIDS, chemotherapy, post-surgery)
- Disseminated infections arise from hematogenous spread from
the primarily infected locus
Candida
- is a yeast and is part of the normal flora (commensal) of the
Skin lesions resulting from skin, mouth, vagina and GI tract.
dissemination from the lungs - most common cause of opportunistic mycoses worldwide
- Candida albicans is the most pathogenic and most commonly
encountered species
Thrush
- a superficial Candida infection of the mouth or vagina

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Oral Thrush - The clinical manifestation and severity of the disease depends
- the white material consists of budding yeast cells and upon the immunologic state of the patient.
pseudohyphae - Lowered host resistance:
 debilitating disease
 neutropenia
 disruption of normal flora
- Almost any organ or system in human body may be involved

CRYPTOCOCCOSIS
Cryptococcus
- is an encapsulated yeast found world-wide; it is found in pigeon
droppings, eucalyptus trees, some fruits and contaminated
milk.

Mucocutaneous Candidiasis
- granulomatous lesions involving the hands

- Cryptococcus neoforman is the only species that is

pathogenic to humans
- The primary port of entry is inhalation.
- The course of the infection is usually subacute or chronic

ASPERGILLOSIS
- is a large spectrum of diseases caused by members of the
genus Aspergillus.
- Colonization of the respiratory tract is common
- The organism can infect the lungs, inner ear, sinuses and,
rarely, the eye of previously healthy persons
- Nosocomial occurence of aspergillosis due to catheters and
other devices is also frequently observed.
- The three principal entities are:
 Invasive Aspergillosis - AIDS is the most commonly encountered predisposing factor
 Allergic Bronchopulmonary Aspergillosis for development of cryptococcosis
 Pulmonary Aspergilloma
- Aspergillus spp. may also be local colonizers in previously
developed lung cavities due to diseases such as
tuberculosis and emphysema (aspergilloma or fungus
ball)

- Cryptococcus is neurotropic and the most common clinical

presentation is meningoencephalitis

ANTIFUNGAL AGENTS
Aspergillus
1. Polyene Antifungal Drugs
- is a filamentous mould and is a ubiquitous fungus found in
- these drugs interact with ergosterol in the fungal cell
nature (soil, plant debris, and indoor air)
membrane and form pores
- second most commonly recovered fungus in opportunistic
 Amphotericin
mycoses (following Candida spp).
 Nystatin
 Pimaricin
Amphotericin B
- first isolated from Streptococcus nodosus in 1955
- an amphoteric compound composed of a hydrophilic
polyhydroxyl chain along one side and a lipophilic polyene
hydrocarbon chain on the other

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- Azoles have five-membered organic rings that contain either
two (imidazole) or three nitrogen molecules (triazoles)
- These agents are thought to inhibit cytochrome P450 14a-
demethylase (P45014DM).
- This enzyme is in the sterol biosynthesis pathway and converts
lanosterol to ergosterol
 Fluconazole
 Itraconazole
- Mechanism of action is the same for all of the preparations and  Ketoconazole
is due to the intrinsic antifungal activity of amphotericin B
- preferentially binds to ergosterol, the primary sterol in fungal
cell membranes.
- this binding disrupts osmotic integrity of the membrane
resulting in the loss of intracellular potassium, magnesium,
sugars and metabolites.
- poorly soluble in water
- must be administered intravenously and is associated with
numerous side effects, which may be severe
- There are several formulations of Amphotericin B.
 Fungizone® (D-AMB)
- is the classic amphotericin B formulation and has been available 3. Allylamine Antifungal Drugs
since 1960 - inhibit squalene epoxidase, a critical enzyme in the
- it is a colloidal suspension of amphotericin B with deoxycholate ergosterol biosynthetic pathway.
(a bile salt) as a solubilizing agent.  Terbinafine
- this preparation has a number of toxicities, which has led to the  Naftifine
development of alternate lipid carrier formulations. Terbinafine
- the major goal of lipid carriers has been to attain a preparation - a synthetic antifungal agent
with lower toxicity but similar efficacy as the deoxycholate - inhibits squalene epoxidas
preparation - this enzyme is part of the fungal sterol biosynthetic pathway
 Amphotericin B colloidal dispersion (ABCD) required to synthesize ergosterol
- a lipid formulation composed of amphotericin B complexed with - mainly effective on dermatophytes (topical or PO)
cholesteryl sulfate
 Amphotericin B lipid complex (ABLC)
- a lipid formulation composed of amphotericin B complexed with
dymyristoyl phosphatidylcholine and dimyristoyl
phosphatidylglycerol
 Liposomal amphotericin B (L-AMB)
- a lipid formulation composed of amphotericin B complexed with Naftifine
hydrogenated soy phosphatidylcholine, - also inhibits squalene epoxidase
distearoylphosphatidylglycerol, and cholesterol - is a topical agent used to treat:
- this preparation is a true liposome composed of unilamellar lipid  athlete's foot (ringworm of the foot; tinea pedis)
vesicles  jock itch (ringworm of the groin; tinea cruris)
Nystatin  ringworm of the body (tinea corporis)
- was the first successful antifungal antibiotic to be developed
and it is still in general use. 4. Morpholine Antifungal Drugs
- The promise of its broad-spectrum antifungal activity is offset - Inhibit the ergosterol biosynthetic pathway at a later step
by host toxicity.  Amorolfine
- It is typically limited to topical use.
5. Antimetabolite Antifungal Drugs
- Flucytosine (5-fluorocytosine) is converted to 5-fluorouracil in
fungal cells, which inhibits DNA, RNA and protein synthesis
Antimetabolites
- Flucytosine (5-fluorocytosine) is an NH2
analong of cytosine
- it is activated by deamination within the
fungal cells to 5-fluorouracil
F
(mammalian cells do not have the
N
Pimaricin (natamycin opthalmic)
- is used topically to treat superficial mycotic infections of the enzyme)
eye. - it is converted to 5-FU-
- It is active against both yeasts and moulds triphosphate, which interferes
O with fungal DNA, RNA and O N
protein synthesis
O OH
O
H
H3C O OH
HOOC

OH
6. Echinocandin Antifungal Drugs
- Presumably inhibit 1,3-β-glucan synthehase
OH - 1,3-β-glucan is required for fungal cell wall biosynthesis
O OH
O
NH2 - Caspofungin
CH3 - Micafungin
2. Azole Antifungal Drugs - Anidulafungin
- inhibit cytochrome P450’s (C14-demethylase) involved in Echinocandins
ergosterol biosynthesis - these agents block the synthesis of a major fungal cell wall
component, 1,3-β-glucan
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- The presumed target is 1,3-β-glucan synthehase - airborne transmission of this low-virulence organism leads to a
dormant, asymptomatic infection
- is a common cause of pneumonia in immunocompromised
patients and is a major cause of opportunistic infections,
morbidity and mortality in AIDS patients
Life Cycle
Three distinct morphological stages:
 Trophozoites (trophic forms)
- uninucleate ameboid-like cells
- this form adheres to alveolar walls and probably multiply by
binary fission.
 Sporocyte
- intermediate between trophozoites and cysts.
 Cyst
- double cell wall
- probably the most immunogenic stage
- Mature cysts contain 6 to 8 intracystic bodies (spores)

7. Miscellaneous Antifungal Drugs

Griseofulvin
- antifungal agent first isolated
from a Penicillium spp. in 1939
- inhibits fungal mitosis by
disrupting the mitotic
spindle through interaction
with polymerized
microtubules
- the drug is insoluble in water
- is mainly effective on
dermatophytes

Transmission
 After inhalation, mature cysts reach the alveoli where they
rupture and release intracystic bodies
 The haploid bodies fuse forming diploid trophozoites that
develop into cysts. (sexual replication)
 Binary fission (asexual replication) of the trophozoites also
occurs, which is thought to be the primary mode of replication
in the lung
 The organism multiplies slowly but extensively in the lungs,
which progressively fills the alveoli with a foamy exudate
consisting of clusters of Pneumocystis jiroveci, degenerated
cells, host proteins, and few alveolar macrophages

Clinical Presentation
 Dyspnea, SOB
 Cyanosis
 Non-productive cough
 Fever
PNEUMOCYSTIS  Chest radiography demonstrates bilateral infiltrates
Pneumocystis jiroveci  Extrapulmonary lesions occur in < 3% of patients
- (formerly P. carinii) was previously considered a protozoan, but o lymph nodes
is in fact a fungus o spleen
- commonly found in the environment and in the lungs of healthy o liver
humans and other animals o bone marrow
- Pneumocystis is a commensal of many animals and human  Increasing pulmonary involvement leads to death in untreated
infection is commonly derived from dogs patients

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Diagnosis
 Clinical Sx
 Chest Radiograph
 Identification of organisms in bronchopulmonary secretions
(sputum/bronchoalveolar lavage)

Treatment
 TMP-SMX, 21 days HIV, 14 days non-HIV
 Dapsone plus trimethoprim
 Pentamidine (inhalation, parenteral)
 Trimetrexate (parenteral)

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