Subcutaneous Fungi MYCO&VIRO

Prof. Benida Fontanilla LE 04

14 February 2022 TRANS 01

OUTLINE
I. Classification of Fungi
A. Zygomycetes
B. Ascomycetes
C. Basidiomycetes
II. Categories of Major Mycoses
III. Subcutaneous Mycoses
A. Sporotrichosis
B. Chromoblastomycosis
C. Mycetoma
D. Phaeohyphomycosis
IV. Rhinosporidium seeberi
V. Lacazia loboi
VI. Entomophthoromycoses
A. Basidiobolus spp
B. Conidiobolus spp
VII. Mucorales
A. Rhizopus spp.
B. Mucor spp.
C. Rhizomucor spp.
D. Lichtheimia spp. (formerly absidia)
E. Saksenaea spp.
I. CLASSIFICATION OF FUNGI
• According to Phyla
A. ZYGOMYCETES
• Sexual reproduction results in zygospores; asexual
reproduction occurs via sporangia; vegetative hyphae are
sparsely septate

Figure 1.

• Under this classification system, the vast majority of human

Figure 1. Sporangiophore, Sporangium, Columella, Sporangiospores, Stolon,
Rhizoid
zygomycotic disease is caused by the members of the family
Mucoraceae.
• Members of this family include zygomycetes that produce
asexual sporangiospores in a sack-like structure called
sporangia.
• A more recently proposed reclassification of the Mucorales by
von Arx places the mucoraceous zygomycetes into seven
families containing human pathogens
• The most common pathogens in this family are in the genera
Rhizopus and Absidia.

• The zygomycetes fall into a distinctive phylum, the phylum

Zygomycota.
• Composed of the organisms that are characterized by the
formation of wide ribbon-like aseptate hyaline hyphae
(coenocytic hyphae) and sexual reproduction with the formation
of zygospores.
• This phylum is divided into two classes, the trichomycetes,
which are obligate symbionts of arthropods and contain no
human pathogens, and the zygomycetes, the class containing
the human pathogens.
• This class is subdivided into two orders, which contain the
agents of human zygomycosis, the mucorales and
entomophthorales

Figure 2.

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C. BASIDIOMYCETES
• Sexual reproduction results in basidiospores supported by
clubshaped basidium; complex hyphae

Figure 6.

Figure 3.

B. ASCOMYCETES
• Sexual reproduction involves a sac or ascus; asexual
reproduction via conidia; molds have septate hyphae

Figure 7.

II. CATEGORIES OF MAJOR MYCOSES

Classification of fungal infections:
• Cutaneous – limited to the epidermis
• Subcutaneous – infection penetrates beneath the skin
• Systemic – infection is deep within the body or disseminated
to internal organs
• Opportunistic – infects individuals who have predisposing
Figure 2. Class Ascomycetes – Ascus and Paraphyses; Class Basidiomycetes – conditions such as an immunodeficiency or debilitating disease
Basidia and Basidiole (Left)
III. SUBCUTANEOUS MYCOSES
• Usually confined to the subcutaneous tissues
• Causative agent – fungi reside in soil or vegetation
• Enter the skin or subcutaneous tissue by traumatic inoculation
with contaminated material
• Lesions become granulomatous and expand slowly from the
area of implantation
• Refer to Figure n
A. SPOROTRICHOSIS
• Caused by Sporothrix schenkii
Figure 3. Ascomycetes (Penicillin) – Mycelium, Hyphae, Spores
• In room temperature: grows as mold with branching septate
hyphae and conidia, delicately clustered at the ends of tapering
conidiophores;
• Colony - blackish and shiny, wrinkled and fuzzy with age

Figure 4.

Figure 8.

• In tissue or in vitro: small budding yeasts

• At 35°C on a rich medium –budding yeasts fusiform in shape

Figure 9.
Figure 5.

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1. CLINICAL FINDINGS
• History of trauma: among agricultural workers; animals
• Multiple subcutaneous nodules and abscesses occur along
lymph nodes
• Granulomatous nodules that progress to form necrotic or
ulcerative lesions

Figure 13. Phialophora verrucose (Left); Fonsacaea pedrosoi (middle);

Cladophialophora carrionii (right)

1. EPIDEMIOLOGY
• Occurs in the tropics
• Fungi are saprophytic; in soil and vegetation
• Among agricultural workers
• Non-communicable
2. CLINICAL FINDINGS
• Fungi are introduced into the skin by trauma, often on legs or
feet
• Over months or years, the primary lesion becomes verrucous
and wart-like with extension along the draining lymphatics
• Cauliflower-like nodules with crusting abscesses eventually
Figure 10. cover the area
2. DIAGNOSIS • Small ulcerations or “black dots”of hemopurulent material on
warty surfaces
• Biopsy material or exudate from lesions
• Examined directly with KOH
• In biopsy: stain with Gomori Methenamine Silver – cell wall
stained black
→ Stain with Periodic Acid-Schiff – cell wall is red

Figure 14.

3. DIAGNOSIS
• Skin scrapings or biopsies from lesions in 10% KOH
• Examine for dark, spherical cells.
• Detection of sclerotic bodies is diagnostic
Figure 11. Asteroid body – yeast cell surrounded by radia2ng deposits of Ag-Ab
• Tissue sections show extensive hyperplasia of the dermal
complexes and complement (lower right) tissue and granulomas
3. TREATMENT
• Some cases self-limited
• Doc: oral itraconazole

Figure 15.

4. TREATMENT
• Surgical excision with wide margins for small lesions
• For larger lesions, chemotherapy with flucytosine or
Figure 12.
itraconazole

B. CHROMOBLASTOMYCOSIS C. MYCETOMA
• Subcutaneous mycotic infection caused by inoculation by • Chronic subcutaneous infection induced by traumatic
fungal agents residing in soil and vegetation inoculation of any of
• Dematiaceous fungi having melaninized cell walls • Several saprophytic species of fungi (eumycetoma)
→ Phialophora verrucosa (maduromycosis; madura foot) or actinomycetous bacteria
→ Fonsacaea pedrosoi (actinomycetoma)
→ Rhinocladiella aquaspersa • Fungal agents:
→ Fonsacaea compacta → Pseudallescheria boydii
→ Cladophialophora carrionii → Madurella mycetomatis

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→ Madurella grisea
→ Exophiala jeanselmei • Refers to infections caused by many kinds of dark, melanin-
→ Acremonium falciforme pigmented dematiaceous fungi
→ Bipolaris,
→ Cladophialophora,
→ Cladosporium,
→ Exophiala,
→ Fonsecaea,
→ Phialophora,
→ Ochronosis,
Figure 16. P. boydii (left); M. mycetomatis (right) → Rhinocladiella, and
1. CLINICAL FINDINGS → Wangiella
• It is distinguished from chromoblastomycosis and mycetoma by
• Subcutaneous tissues of the feet, lower extremities, hands and
the absence of specific histopathologic findings
exposed areas
• Although some species of these fungi may be true pathogens
• Local swelling and interconnecting, draining sinuses containing
and cause phaeohyphomycosis in immunocompetent patients,
granules
pigmented fungi have been increasingly recognized as
• Untreated lesions persist for years and extend deeper and
opportunists; almost all cases of widely disseminated infection
peripherally
occur in immunosuppressed patients
• Can have deformation and loss of function
• Dematiaceous fungi only rarely cause fatal infections in
patients who have intact host defense mechanisms,
• Clinical syndromes include:
→ Invasive sinusitis, sometimes with bone necrosis
→ Subcutaneous nodule or abscesses
→ Keratitis
→ Lung masses
→ Osteomyelitis
→ Mycotic arthritis
→ Endocarditis
→ Brain abscess, and
Figure 17. → Disseminated infection
2. DIAGNOSIS 1. DIAGNOSIS
• Pus or biopsy material for examination and culture • Examination using Masson-Fontana staining
• The granule color, size and presence of hyaline or pigmented • Culture to identify causative species
hyphae are helpful • Dematiaceous fungi can frequently be discerned in tissue
• In determining the causative agent specimens stained with conventional hematoxylin and eosin
→ They appear as septate, brownish hyphae or yeast-like
cells, reflecting their high melanin content
→ Masson-Fontana staining for melanin confirms their
presence
→ Phaeohyphomycosis is distinguished from
chromoblastomycosis and mycetoma by the absence of
specific histopathologic findings such as sclerotic
Figure 18. Granule (left); Hyphae at periphery of granule (right)
bodies or grains in tissue
3. TREATMENT • Culture is needed to identify the causative species
• surgical debriment or excision and long periods of a. BIPOLARIS
chemotherapy to penetrate the lesions
• Kingdom: Fungi
• Pseudallescheria boydii – topical nystatin or miconazole
• Phylum: Ascomycota
• Madurella infection – Itraconazole, ketoconazole and
• Class: Euascomycetes
amphotericin B
• Order: Pleosporales
• Exophiala jeanselmei – flucytosine
• Family: Pleosporaceae
D. PHAEOHYPHOMYCOSIS • Genus: Bipolaris
• Term applied to infections characterized by the presence of
darkly pigmented septate hyphae (Exophiala spp, Phialophora
spp.)
• Clinical presentations: solitary encapsulated cysts in the
subcutaneous tissue, sinusitis, brain abscess
• In tissue, hyphae are large with yeast cells with melanin in cell
walls
Figure 20.

Figure 21
Figure 19. Brown hyphae in brain tissue (left); Immunocompromised child (right)
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A. Ellipsoid conidia with the common number of 3-5 • Cultured Fonsecaea pedrosoi colonies are slow growing,
septations stained with lactophenol cotton blue lanose to velvety, olivaceous to black, which present growth
counterstain consistently at temperatures up to 35oC for the clinical isolates,
B. a cluster of conidia surrounded by less lactophenol cotton whereas the environment isolates exhibit growth consistently
blue better demonstrating brown melanin pigment in the up to 35oC, and irregularly up to 37oC
cell wall, and • In microscopy, Fonsecaea pedrosoi colonies are characterized
C. a dark gray speckled fungal colony by dark hyphae and suberect conidiospores loosely
b. CLADOPHIALOPHORA branched
• Kingdom: Fungi d. CLADOSPORIUM
• Phylum: Ascomycota • Conidiospores and conidial chains
• Subphylum: Ascomycotina
• Genus: Cladophialophora

Figure 22.
Figure 26

Figure 23.
A. KOH (20%) mount showing dematiaceous hyphae and Figure 27. Cladosporium cladosporioides
yeast-like cells • Colonies are rather slow growing, mostly olivaceous-brown to
B. Histopathological examination revealing septate blackish brown but also sometimes grey, buff or brown suede-
dematiaceous hyphae (400x) like floccose, often becoming powdery due to the production of
C. Macromorphological features of Cladophialophora abundant conidia
bantiana on PDA medium showing a velvety, grayish
black, and olive green colony after 20 days growth IV. RHINOSPORIDIUM SEEBERI
D. Microscopic morphology of C. bantiana from Riddell • Causes Rhinosporidiosis
culture, showing the long chains of ellipsoidal conidia and → which is manifested as tumor-like polyps developing
poorly differentiated conidiophores (400x) primarily in the nostrils and conjunctiva in human and
animals.
c. FONSECAEA
• Clinical features: depends on the site of involvement.
• Kingdom: Fungi → usually seen in nasal cavity and nasopharynx.
• Phylum: Ascomycota → In the nasal cavity the lesions begin as sessile masses
• Class: Euascomycetes → gradually grow to become fleshy pedunculated polypoid
• Order: Chaetothyriales mass.
• Family: Herpotrichiellaceae • Gross: These are reddish polypoidal, bulky, friable mucosal
• Genus: Fonsecaea masses.
• R. seeberi was first reported in 1900 as a sporozoan parasite,
but later classified as a lower fungi,

Figure 24

Figure 28. (Left) R. seeberi in nasal cavity. (Right) R. seeberi in nasopharynx

• presence of large, round-shaped mature stage and small

endospores with resistance to culturing.
• Mature sporangia are 100 to 450 micrometers in diameter
with a thick chitinous wall and contain sporangiospores in
different stages of development.
Figure 25 • Sporangiospores stain basophilic and measures 7 to 9 microns
in diameter.

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• If there is rupture of sporangia in the stroma, an intense
granulomatous response can be seen.
Figure 29
Figure 30. (Left) actual biopsy showing very large sporangium with
sporangiospores
Figure 31. Electron microscope tissue section showing a Rhinosporidium
seeberi. Mature endoconidia are in the middle while immature endoconidia is in
the periphery
• Electron microscopy tissue section showing a Rhinosporidium
seeberi mature sporangium containing hundreds of
endoconidia, one already outside the sporangium and the
others in the process of being expelled through a pore (arrow
heads) (a video depicting the endoconidia release is available
at: http://www.bld.msu.edu/Rhino).
• Note the presence of a thin cell wall and the formation of three
prominent inner layers (a clear space between the mature
endoconidia and the cell wall) primarily located near the pore.
• At this magnification this structure appears as a single inner
layer, but it comprises three well defined inner layers.
• The presence of fully developed endoconidia is observed at
the center and toward the pore, whereas immature small
usually oval endoconidia are found at the opposite site (white
heads), a distinctive feature of mature sporangia.
• Three immature sporangia are also noted in the lower section
of the mature sporangium (white arrow heads) (Bar=100(μm).
Figure 32
A. Rhinosporidiosis. Clinical image showing a ‘strawberry
mass’ just behind the uvula
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B. Excised specimen
C. Histopathology showing multiple spores in different stages
of maturation (black arrows) along with sporangiospores
(white arrows) suggestive of rhinosporidiosis (H&E ×40)
Figure 33
• Rhinosporidium seeberi, a microorganism that can infect the
mucosal surfaces of humans and animals, has been classified
as a fungus on the basis of morphologic and histochemical
characteristics.
• Using consensus polymerase chain reaction (PCR), we
amplified a portion of the R. seeberi 18S rRNA gene directly
from the infected tissue.
• Analysis of the aligned sequence and interference of the
phylogenic relationships showed that R. seeberi is a protist from
a novel clade of parasites that infect fish and amphibians.
• Flourescence in site hybridization and R. seeberi-specific PCR
showed that this unique 18S rRNA sequence is also present in
other tissues infected with R. seeberi. Our data support that the
R. seeberi phylogeny recently suggested by another group.
• R. seeberi is not a classic fungus, but rather the first known
human pathogen from the DRIPs clade, a novel clade of
aquatic prostistan parasites (Ichthyosporea).
V. LACAZIA LOBOI
• has been known by various names such as:
→ Blastomyces brasiliensis,
→ Blastomyces loboi,
→ Glenosporella loboi,
→ Glenosporopsis amazonica,
→ Loboa loboi,
→ Lobomyces loboi, and
→ Paracoccidioides loboi.
• Herr et al reported that L. loboi is phylogenetically linked to P.
brasiliensis and to the other dimorphic fungal Onygenales.
• The genus Lacazia contains a single species, Lacazia loboi.
• While the name Loboa loboi is still frequently used to refer to
the causative agent of lobomycosis, more recently,
classification of the fungus in the genus Lacazia and
conclusively, the name Lacazia loboi has been proposed by
McGinnis et al
Figure 34
• Kingdom: Fungi
• Phylum: Zygomycota
• Subphylum: Zygomyco/na
• Order: Entomophthorales
• Family: Uncertain
• Genus: Lacazia
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VI. ENTOMOPHTHOROMYCOSES
• Entomophthoromycoses, formally classified as a subgroup of
phycomycoses and, later, zygomycoses, are rare, invasive
fungal infections characterized by the formation of solid
tumefactions.
• Diseases due to entomophthoralean fungi are endemic in
regions of tropical (rhino- and subcutaneous
Figure 35 entomophthoromycosis) and arid (gastrointestinal
entomophthoromycosis) climates

Figure 36. A) Multiple, confluent, keloid-like, hypochromic nodules with flat shiny
surfaces involving the entire free border, posterior aspect, and lobule of the left
ear of a fisherman, Venezuela. B) Numerous Lacazia loboi tissue-phase
organisms within the stroma. Note the typical chain pattern showing simple
gemation budding (Gomori-Grocott stain, magnification x100). C) Yeast cells
showing typical double refraction of the membrane and protoplasmic bodies
within cells (periodic acid-Schiff stain, magnification x600).
• Numerous yeast-like, round, thick-walled cells are visualized.
Chains of yeast cells are typically formed. Little tube-like
connections are visible between the yeast cells
• The etiologic agent of Jorge Lobo's disease (lacaziosis),
Lacazia loboi is an uncultivated fungal pathogen of humans and
dolphins causing cutaneous and subcutaneous infections
and, rarely, visceral involvement.
• This anomalous pathogen is restricted to Mexico, Central
America, and South America.
• Cases in dolphins in the coasts of France and the USA with
transmission to aquarium personnel have also been Figure 40. Human Pathogenic Entomophthorales
documented.
• Entomophthoralean fungi (Conidiobolus coronatus,
• The first case was diagnosed in 1930 by Jorge Lobo in a Conidiobolus incongruus, Conidiobolus lamprauges, and
Brazilian human patient with cutaneous parakeloidal lesions. Basidiobolus ranarum) live as saprophytes in soil and decaying
• Since then, hundreds of new cases have been recorded in Latin plant matter.
America, but Brazil has the highest incidence. • Species were also isolated from surface water or feces of
insectivores.
• Their ability to destroy insects coined the name of
Entomophthorales
• These fungi cause infections in humans and mammals (e.g.,
horses, sheep, dogs, chimpanzees, and llamas).
• Potential sources of infection are contaminated soil, leaf litter,
insects, and water

Figure 37. Keloid-like lesions and diffuse infiltration on the earlobe and posterior
helix
Figure 38. Acanthosis of the epidermis. All through the dermis, amidst a fibrous
stroma, a large number of round structures with mild interspersed
inflammatory lymphocytic infiltrate can be seen (Hematoxylin & eosin, X100)
A. BASIDIOBOLUS SPP.
• Fungi from the order Entomophthorales are rare but well
recognized cause of tropical fungal infection
• typically causing subcutaneous truncal or limb lesions in
immunocompetent hosts.
• may also mimic malignancy by causing intrabdominal mass,
sometimes resulting in obstructive gastrointestinal or renal
presentations.
• Basidiobolus spp is an unusual cause of infection with
characteristic mycological and histopathological findings.
• Infection can present in a number of ways ranging from a slow-
growing mass in the subcutaneous soft tissue to an invasive
mass in the gastrointestinal tract.

Figure 39. Chain formation, isolated and budding fungi (Hematoxylin & eosin, Figure 41. The life cycle of Basidiobolus ranarum
X400)

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A The life cycle starts when sticky conidia are forcibly

ejected from sporangiospores
B The sticky primary conidium could attach to a passing
host (humans or insects) or develop an elongated
adhesive conidium (capilloconidium), which also can
attach to passing hosts
C The latter secondary elongated structure could
develop to contain a sticky beak haptor that divides Figure 43
to form numerous “Palmella” endospores, some of
which are released outside the broken
capilloconidium cell wall, giving rise to new hyphae
and single sporangiophores (A)
D and E The target insects (D) can be ingested by reptiles or
amphibians (E), initiating a new cycle inside the
intestinal tract of these animals
F In this new environment, hundreds of resistant
meristospores are produced and then secreted in
feces
G When environmental conditions are right, coenocytic Figure 44. (Left) zygospore with characteristic beaked appendage. (Middle)
Beaked-like appendage characteristic of Basidiobolus; Zygospores. (Right) thick
hyphae are developed walled zygospore
H If two opposite-sec hyphae contact each other, their
exchange of genetic material leads to the formation
of sexual sygospores. Zygospores can develop into
sporangiospores (long arrow)

• Identification of its unique beak-like zygospore and

Splendore-Hoeppli phenomenon on histopathological
specimens can be pathognomonic and could provide the key
to early diagnosis.

Figure 45.

A Creamy rugose colony of Basidiobolus ranarum on

Figure 42.
A. Periodic acid-Schiff stain showing a dermal giant cell (long
arrow) with a short, irregular, broad pauci-septate hyphal
remnant (short arrow).
B. No stain used. Zygospore of Basidiobolus ranarum
measuring 25mm in diameter, with two break-like
appendages. Large vegetative hyphae, 8mm in diameter, are
seen on the left (arrow).
C. Yellowish-grey, radially folded Basidiobolus ranarum
colonies after 7 days of incubation at 308C on Sabouraud’s
dextrose agar, placed upon a black background. Satellite
colonies are seen (arrow).
D. Lactophenol cotton blue stain demonstrating a sporosphore
with a sub-sporangial vesicle on the left and a conidia on the
right
1. MICROSCOPIC FEATURES
• The hyphae of Basidiobolus are large (8 to 20 µm in diameter)
and septate.
• Having large hyphae/thick fungal walls are a common
characteristic of fungi in the phylum Zygomycota.
• The more spores the fungus produces, the greater the number
of septa are present.
• Basidiobolus spp. produce sexual spores called zygospores
that are thick-walled and can be smooth or have undulating
outer cell walls.
• This fungus can also produce two different types of asexual
spores or conidia called: ballistospores and capilliconidia
2% Sabouraud dextrose agar.
B to E Encounter of opposite-sex hyphae before the
formation of lateral beaks. Bars, 12 m (B to D) and 10
m (E).
F and G After the exchange of genetic material, zygospores
develop with their characteristic beak. Bars, 25 m (F)
and 15 m (G)
2. MYCOLOGICAL AND CLINICAL ASPECTS OF
Basidiobolus ranarum
• A culture of B. ranarum on 2% Sabouraud dextrose agar after
4 d incubation at 37 ̊C. Note the small colonies formed by
forcibly ejected conidia growing around the edges of the culture.
• Coenocytic hypha of B. ranarum forming the first septum and
the elongation across the plane in which the future zygospore
will develop.
• Three mature zygospores with thick walls and their prominent
beaks.
• Note the Splendore –Hoeppli phenomenon around the empty,
round hyphal structures in a histological section ( D , arrows).
• Single replicative conidium formation in Conidiobolus
coronatus.
• C. coronatus conidia with a typical basal papilla, which marks
the residual section formed after ejection. The inset of panel F
shows a villose conidium.
• C. coronatus multiplicative secondary conidia around a single
conidium. The left portion of panel G shows a sporangiophore
before conidium discharge. The inset depicts two multiplicative
conidia.

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• The eosinophilic inflammatory response and longitudinal
sectioned hyphae of C. coronatus surrounded by an
eosinophilic reaction. The presence of numerous eosinophils
(arrows) is noted.
• A young boy with bilateral rhinofacial infection caused by C.
coronatus .
• Note the scar of the biopsy performed for diagnostic purposes.
Figure 46. Mycological and Clinical Aspects of Basiodiobolus ranarum
• Splendore-Hoeppli phenomenon (asteroid bodies) is the in vivo
formation of intensely eosinophilic material (radiate, star-like,
asteroid or club-shaped configurations) around microorganisms
(fungi, bacteria and parasites) or biologically inert substances.
• The Splendore-Hoeppli reaction material comprises antigen-
antibody complex, tissue debris and fibrin.
• Although the exact nature of this reaction is unknown, it is
thought to be a localized immunological response to an antigen-
antibody precipitate related to fungi, parasites, bacteria or inert
materials.
• The characteristic formation of the peri bacterial or perifungal
Splendore-Hoeppli reaction probably prevents phagocytosis
and intracellular killing of the insulting agent leading to
chronicity of infection.
Figure 47.
NICE-TO-KNOW
• (A) The eosinophilic reaction around an invading hypha
encases basidiobolomycosis or conidiobolomycosis. Note
the presence of IgE, the degranulation of eosinophils around
the hyphae, and several pyknotic nuclei within degranulate
eosinophilic material.
• (B and C) Histopathological sections from a human case of
intestinal basidiobolomycosis. Note the presence of
numerous eosinophils surrounding the cross sections of
hyphae that appear as spherical or oval structures in the
center of the Splendore-Höeppli phenomenon. Under this
perspective, the released IgE (red immunoglobulins) will
bind to cell wall antigens that attract eosinophils to the site
of infection.
• Panels A and B display the binding and degranulation of the
eosinophils on the hyphae triggered by IgE (panel A, red
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immunoglobulins). The eosinophil nuclei at this stage appear
at the periphery of the eosinophilic precipitate (A and B).
• As the lesion becomes old (chronic stages), other
eosinophils will bind the complex, and after degranulation,
their nuclei are also incorporated into the eosinophilic
material and become pyknotic, giving rise to the Splendore-
Höeppli phenomenon (C). In some instances, only the
eosinophilic material will be expressed around the invading
microbe (115, 137).
Figure 48. As highlighted in Fig. 9, the Th2 subset will trigger the release of
IL-4, IL-5, and IL-13, and B cells will express IgE (red immunoglobulins).
B. CONIDIOBOLUS SPP.
1. INFECTIONS CAUSED
• by Conidiobolus coronatus (rhinoentomophthoromycosis) lead
to a chronic, localized subcutaneous infection affecting tissues
of the nose, cheek, and upper lip.
• Conidiobolus coronatus is a mucormycete found in tropical rain
forest areas.
• In contrast to zygomycosis caused by Basidiobolus,
rhinoentomophthoromycosis is a disease of young adults,
rather than children.
• Males are more commonly affected than females.
• Most infections are reported from West Africa, particularly
Nigeria, but cases have been recognized in India and South
America.
• The mode of infection is unknown, but is probably inoculation
of contaminated soil or vegetable matter through minor trauma
or insect bites.
2. CLINICAL MANIFESTATIONS
• Infection apparently originates in the nasal mucosa, leading
to nasal obstruction, which may be unilateral.
• Tissue swelling becomes pronounced affecting the nose and
nasolabial folds, cheeks, and upper lip, eventually producing
gross facial distortion.
• The infected areas have distinct margins but the mass is not
movable over the underlying tissues. There are few symptoms.
• The clinical features are distinctive, i.e. localized swelling of
the nose and face in tropical countries.
• Biopsy may be diagnostic when characteristic hyphae are
present and associated with an eosinophilic sheath and
eosinophilic granuloma, same as with Basidiobolus infections.
• Conidiobolus and Basidobolus infections can be differentiated
by the site involved.
Figure 49. Clinical manifestation of Conidiobolus spp. (Left). Conidiobolus on a
microscope (right).
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• Conidiobolomycosis is an uncommon infectious disease of
tropical areas and is known to cause chronic cutaneous and
subcutaneous granulomatous and systemic infectious diseases
in immunocompetent humans or animals.
• Conidiobolus species are considered saprobes distributed in
vegetation and soil
NICE-TO-KNOW
HUMAN PATHOGENIC ENTOMOPHTHORALES
Refer to Figure 50.
• Putative events based on histopathological and
immunological findings during entomophthoramycosis and
during parasitic infections (112, 133, 137). Under this
scenario, through open skin, a conidium attaches to the host
and produces a germ tube penetrating the host.
• The invading hyphae then release secretory immunogens
(pathogen- associated molecular patterns [PAMPs]).
Dendritic cells (DC), through pattern recognition receptors
(PRRs), contact the antigen and are activated, becoming
antigen-presenting cells (APCs). The activated APCs
process the antigens and release IL-4 during migration to
nearby lymph nodes to present the antigen to Th0 naive
cells.
• The Th0 naive cells in turn release IL-4 and IL-10 and
become a powerful Th2 subset. The Th2 subset releases
more IL-4, IL-5, IL-13, and IL-10, resulting in the
downregulation of Th17 and Th1 subsets.
• These interleukins activate the differentiation of alternative
activated macrophages (AAM) that could inhibit the
proliferation of cells such as Th1, Th2, and Th17 cells.
However, the exacerbated production of IL-4 and IL-5 by the
Th2 subset drives the immune response into a strong Th2
subset.
• In turn, IL-4 and IL-13 stimulate B cells to produce precipitin
IgG (detected by serological assays in cases of
entomophthoramycosis) and IgE as well as the activation of
effector cells such as mast cells, eosinophils (EO), and
basophils. The released IgE will also specifically bind to the
invading hyphae, and the eosinophils will in turn attach to the
Fc region of IgE, triggering the degranulation of the
eosinophils around the invading hyphae. A similar outcome
occurs after IgE binding to mast cells and basophils, causing
fibrosis and tissue damage, consistent with the clinical
features of entomophthoramycosis. TGF-, transforming
growth factor.
Figure 50. Human Pathogenic Entomopthorales
MV
VII. MUCORALES
• The traditional Zygomycota have undergone taxonomic
changes. Taxa traditionally placed in the phylum Zygomycota
are now assigned among the phylum Glomeromycota and four
subphyla, Entomophthoromycotina, Mucoromycotina,
Kickxellomycotina, and Zoopagomycotina.
• Mucorales, the most clinically significant order, generally
produce profuse, gray-to-white, aerial mycelia characterized by
the presence of hyaline, sparsely septate hyphae. Asexual
reproduc- tion is characterized by the presence of
sporangiophores and sporangiospores. The asexual spores
(sporangiospores) are produced in a structure known as a
sporangium, which develops from a supporting structure
termed a sporangiophore
• Although some Mucorales are capable of sexual reproduction
resulting in the production of zygospores, these structures are
not routinely seen in clinical laboratories.
*from book
A. RHIZOPUS SPP.
• Rhizopus is probably the best-known genus in the class of
Zygomycetes fungi, which normally live on dead and decaying
plant material.
• These fungi exhibit a complex metabolism and produce a
variety of enzymes that enable them to utilize a wide range of
nutrients.
• Several different strains of Rhizopus are used to produce
fermented foods and beverages, particularly in Southeast Asia.
• In addition, Rhizopus has several industrial applications in the
manufacturing of enzymes, including amylases, pectinases,
cell upases, proteases, and phytases, and metabolites such as
ethanol, lactic, and fumaric acids. The cell mass of these fungi
also has found applications in food and feed industries.
Figure 51.
• Differs from the molds previously described, because it is
nonseptate and produces sporangiospores rather than conidia.
• It is a very-fast-growing, spreading type of mold which has white
mycelia and black sporangia.
• Rhizopus species form rhizoids at the base of the
sporangiophores, and columella in the sporangium (Figure 6).
• Young sporangia are white before turning black with age.
Probably the most common of the Rhizopus species is R.
stolonifer, the so-called bread mold. Besides bread, Rhizopus
causes spoilage of strawberries, other berries, fruits, and
vegetables.
• Rhizopus species have also been isolated from cereal grains,
nuts, and meat. R. oligosporus is used in making tempeh and
certain other mold-fermented foods.
• Rhizopus is placed in the taxonomic order Mucorales and is
somewhat typical, and probably the most common genus of that
order to contaminate foods.
• Rhizopus organisms have an enzyme, ketone reductase, which
allows them to thrive in high glucose, acidic conditions. Serum
from healthy individuals inhibits growth of Rhizopus, whereas
serum from individuals in diabetic ketoacidosis stimulates
growth
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Figure 52
Figure 53. Mature sporangium releasing sporangiospores
Figure 54. Sporangium with Sporangiospores at end of Sporangiophore (left).
Sporangium releasing sporangiospores (Top Right). Sporangiospores (bottom
right)
• Rhino-orbital-cerebral and pulmonary mucormycosis are
acquired by the inhalation of spores.
• In healthy individuals, cilia transport these spores to the
pharynx, and they are cleared through the gastrointestinal tract.
• In susceptible individuals, infection usually begins in the nasal
turbinates or the alveoli.
• The agents of mucormycosis are Angio invasive; thus,
infarction of infected tissues is a hallmark of invasive disease.
Figure 55.
Figure 56.
MV
• Refer to the figure above.
• (Left photo) A 69-year-old man presented (day 0) with a 4-day
history of a skin lesion on his right lower leg, which was initially
erythematous, became black and necrotic, then broke down
into an ulcer. He had no systemic symptoms and could not
recall any trauma or injury to the leg. His history included poorly
controlled type 2 diabetes mellitus, metasta2c nonsmall cell
lung cancer for which he had received chemotherapy and
radiotherapy, chronic obstructive pulmonary disease, recurrent
pulmonary emboli, peripheral vascular disease, and chronic
renal impairment. He had received large doses of prednisolone
for radiation pneumonitis and exacerbations of his airways
disease and had previously been a heavy smoker.
• (Upper right) Biopsy of ulcer performed on day 4: H&E stain,
400× (a) and PAS stain, 400× (b) showing narrow-angled
branching fungal hyphae invading into necrotic dermis.
• (Bottom right) Sabouraud agar plate following 24 h incubation
(a) and view from slide culture (40×) showing the presence and
orientation of rhizoids (b).
B. MUCOR SPP.
• The Mucor fungi cause the group of infections referred to as
zygomycosis (mucormycosis).
• Mucor is a filamentous fungus found in soil, plants, and
decaying fruits.
• The genus has several species, the more common ones being
→ Mucor amphibiorum,
→ M. circinelloides,
→ M. hiemalis,
→ M. indicus,
→ M. racemosus, and
→ M. ramosissimus.
• Zygomycosis includes mucocutaneous and rhinocerebral
infections as well as septic arthritis, dialysis-associated
peritonitis, renal infections, gastritis, and pulmonary infections.
• Clinical complications arise as a result of vascular invasion
that causes necrosis of the infected tissue and to
perineural invasion. Morphologically, the organisms are seen
as hyphae, sporangiophores, sporangia, and spores.
Figure 57. A photomicrograph depicting a mature sporangium exhibited by a
Mucor sp. fungus. Mucor spp. are common indoor molds, and is among the fungi
that cause the group of infections known as zygomycosis. The infection typically
involves the rhino-facial-cranial area, lungs, GI tract, skin, or less commonly other
organ systems.
Figure 58. Rhinocerebral Mucormycosis
NEED-TO-KNOW
• Mucormycosis is an infection of immunosuppressed
patients, including those with diabetes, In rhinocerebral
mucormycosis, Rhizopus, Rhizomucor, or another
angioinvasive fungal species enters the vascular space and
causes tissue necrosis of the nasal septum. Palate, orbit,
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and sinuses. The infection can extend into the brain, causing
cavernous sinus thrombosis, seizures, and thrombotic
stroke.
1. DIAGNOSIS OF MUCORMYCOSIS
• Examination of tissue samples for broad, ribbon -like,
nonseptate hyphae.
2. CULTURE
• Diagnosis of mucormycosis requires a high index of suspicion
and painstaking examination of tissue samples for large
nonseptate hyphae with irregular diameters and right-angle
branching patterns; the examination must be thorough because
much of the necrotic debris contains no organisms.
• For unclear reasons, cultures may be negative, even when
hyphae are clearly visible in tissues.
• CT and x -rays often underestimate or miss significant bone
destruction.
controlled diabetes, metabolic acidosis, steroid therapy, solid
organ and hematopoietic stem cell transplant, penetrating
trauma, burns, neutropenia, iron overload, and deferoxamine
therapy.
• Many centers have seen a rise in incidence of opportunistic
infection in neutropenic patients over the past 10 years with
increasing use of voriconazole prophylaxis and changing
chemotherapeutic regimens.
• Locally invasive disease most commonly involves the lungs
and/or sinuses but may involve any organ, including the GI tract
or skin.
• Disseminated disease may develop from locally invasive
disease that spreads hematogenous.
• Cases have also been reported related to intravenous (IV) drug
abuse and peritoneal dialysis catheters.
• After dissemination, any organ may be affected.
•

Figure 59. Sporangia, columella and sporangiospores of Mucor spp.

Figure 61. Rhizomucor variabilis var. regularior

(A and B) Macroscopic morphology on potato dextrose agar.

Colonies were filling the entire plate in 5 days at 30°C (A) and
showed restricted growth at 37°C (B). Colonies varied in color from
brown to tan and were hairy, with reverse buff-to-brown color. (C)
Microscopic morphology in lactophenol cotton blue stain after 3 days
(magnification, ϫ 200): hyaline, unbranched, ribbon-like hyphae;
long, simple sporangiophores arising from hyphae and ending
in sporangium; spherical sporangia with globose columella and
no apophysis; hyaline, ellipsoidal, smooth-walled
sporangiospores. The inset shows details of the spherical
columella (arrow) with sporangiospores (magnification, ϫ 400)

Figure 60. Morphology of a typical member species (exemplarily: Mucor flavus) D. LICHTHEIMIA SPP. (FORMERLY ABSIDIA)
of the genus Mucor representing the type of the order Mucorales.
• According to the revised taxonomy, fungi causing
NICE TO KNOW mucormycosis are classified in the (new) phylum
Refer to Figure 60. Glomeromycota, class Glomeromycetes, subphylum
• A: Mature sporangium. Mucoromycotina, order Mucorales.
• B: Mature sporangium with prominently visible columella, a • The genera of Rhizopus, Mucor, Lichtheimia (formerly Absidia),
sterile and bulbous vesicle on the sporangiophore apex. Cunninghamella, Rhizomucor, Apophysomyces, and
• C: Sporangiospores. Saksenaea constitute those that are identified as causative
• D: Immature sporangium. agents of the majority of cases of mucormycosis.
• Approximately half of all mucormycosis cases are caused by
• E: Egg-shaped columella.
Rhizopus spp.
F: Typical appearance of mycelial lawn consisting of well-
developed hyphae after 10 days growth on 3% malt extract agar • Lichtheimia corymbifera is the second and Mucor spp. are the
at room temperature. third most common mucoralean fungi shown to be responsible
for development of mucormycosis
C. RHIZOMUCOR SPP. • The fungi classified in the order Mucorales are mainly
• the zygomycetes are a class of fungi known to cause saprophytic rapidly growing, and able to grow at temperatures
cutaneous, locally invasive, and disseminated infection. higher than 37°C (except for Mucor spp.).
• This class includes • These molds are supposedly ubiquitous in nature and widely
→ Rhizopus spp., found on organic substrates, including bread, decaying fruits,
→ Rhizomucor spp., vegetable matter, crop debris, soil between growing seasons,
→ Absidia spp., compost piles, and animal excreta.
→ Apophysomyces spp., • The existence of non-septate (or pauciseptate) hyphae is
typical and responsible for the rapid growth as well as the fragile
→ Cunninghamella spp., and
structure.
→ Mucor spp.
• While the terms mucormycosis, phycomycosis, and
• Infections are rarely seen in normal hosts, occurring almost
zygomycosis have so far been used to refer to the diseases
exclusively in hosts with well-defined risk factors such as poorly

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 4.01 Subcutaneous Fungi

caused by the order Mucorales, the recently revised and • Sporangia are typically flask-shaped with a distinct spherical
accepted term remains as mucormycosis. venter and long-neck, arising singly or in pairs from
dichotomously branched, darkly pigmented rhizoids.
• Collumellae are prominent and dome-shaped.
Sporangiospores are small, oblong, 1-2 x 3-4 µm, and are
discharged through the neck following the dissolution of an
apical mucilaginous plug.

Figure 62. Micro-morphology of Absidia corymbifera (Lacto-phenol cotton blue

staining; original magnification )400 (a) Branching sporangiophores arising from
the stolons )800, (b) pear-shaped sporangia and conical columellae )1200, (c)
flask-shaped apophysis beneath the sporangium )800, (d) branching
sporangiophores arranged in whorls.

Figure 67.

Figure 63. Young slide culture Absidia corymbifera X100 LPCB (Left). Absidia
corymbifera sporangium filled with sporangiospores. (LPCB adhesive tape
prepara2on X400 + additional 1% digital magnification (Middle). Apophysis
structure can be seen in dissolving sporangium on left. (LPCB X400) (Right).
E. SAKSENAEA SPP.
• It is an emerging human pathogen (Holland, 1997) that is most
often associated with cutaneous or subcutaneous lesions after
trauma.
Figure 64. (A) S. erythrospora on a CZA after 3 weeks of incubation at 25C. (B)
Fruiting structure of S. erythrospora depicting lateral rhizoids. A brown, straight,
encrusted sporangiophore; the columella at the base of the sporangium; and the
sporangium with its long neck filled with sporangiospores. (C) Higher
magnification of the neck of the sporangium filled with sporangiospores. (D)
Ellipsoidal, biconcave, sporangiospores.
NEED-TO-KNOW
Key Concepts
1. Subcutaneous mycoses may be caused by dozens of
environmental molds associated with vegetation and soil
2. These infections are usually acquired when minor cuts or
scratches introduce soil or plant debris (eg splinters, thorns)
containing the pathogenic fungi
3. Sporothrix schenckii, the cause of sporotrichosis, is a
dimorphic fungus that converts from hyphal growth to yeast
cells within the host.
4. The diagnostic feature of chromoblastomycosis is the
microscopic observation of brownish (melanized) spherical
sclerotic bodies within the lesions
5. The diagnostic feature of phaeohyphomycosis is the
presence of brownish melanized, septate hyphae within the
lesions
6. The hallmark of mycetoma is localized swelling and the
formation of fistulae that contain hard granules composed of
hyphae and inflammatory tissue (eg macrophages, fibrin)

Figure 65. Histopathology of a section of facial tissue showing broad, aseptate

hyphae

• Morphological description: Colonies are fast growing,

downy, white with no reverse pigment, and made up of broad,
non-septate hyphae typical of a mucoromycetes fungus.

Figure 66. Saksenaea spp. 48 hrs growth on SAB or SDA at 30C

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Figure 68.

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