LPCB Mount Footages of Various Fungi: Introduction, List of Footages and Decriptions

LPCB Mount Footages of Various Fungi: Introduction, List of Footages and Decriptions

Introduction of LPCB Mount Footages of Various Fungi

LPCB Mount Footages of Various Fungi is the collection of fungal elements encountered during LPCB mount microscopy of a variety of clinical specimens. LPCB stain stands for lactophenol cotton blue and it is a combination of fixative, staining, and clearing agent. LPCB uses both as a mounting fluid and a stain. This is used for staining and microscopic identification of fungi. Its contents functions are as follows- Lactic acid: It helps in preserving the morphology of the fungal elements. Phenol: It acts as a disinfectant. Cotton blue: It stains the fungal elements as well as intestinal parasitic (cyst, ova, and oocyst) and non-parasitic structures (vegetable cells, mucus, muscle fibers, and other artifacts). Glycerol: It is a hygroscopic agent that prevents drying

List of LPCB Mount Footages of Various Fungi 

  1. Candida
  2. Cryptococcus
  3. Aspergillus
  4. Mucor
  5. Trichophyton
  6. Microsporum
  7. Epidermophyton
  8. Geotrichum
  9. Malassezia
  10. Bipolaris
  11. Cladosporium
  12. Curvularia
  13. Fonsecaea
  14. Fusarium oxysporum
  15. Fusarium solani
  16. Penicillium
  17. Rhizopus
  18. Scedosporium
  19. Sporothrix schenckii
  20. Syncephalastrum
  21. Trichosporon
  22. Acremonium
  23. Ochroconis gallopava
  24. Trichoderma species

 

LPCB Mount Footages of Various Fungi with Descriptions

 

Candida albicans in LPCB tease mount

 

 

Cryptococcus neoformans in LPCB tease mount

 

 

Aspergillus species in LPCB mount

LPCB Mount Footages

 

 

Aspergillus fumigatus Colony on SDA, LPCB tease mount under microscopy

LPCB Mount Footages

 

 

 

Aspergillus fumigatus growth on SDA and its LPCB Preparation showing conidia, conidiophores, phialides

LPCB Mount Footages

Mucor under the Microscope
Mucor structures under the microscope
Sporangium
Sporangiospores
Columella
Sporangiophore/aerial hyphae
Lacking rhizoids and stolons

 

 

 

 

Mucor with
-aseptate hyphae
– no evidence of root
-presence of sporangium.

 

Mucor in LPCB Preparation

 

 

Mucor causing Mucormycosis colony morphology and LPCB Tease Mount Microscopy

Mucor colony morphology and LPCB Tease Mount Microscopy
Mucormycosis: Introduction, Pathogenicity, Laboratory Diagnosis and Treatment@https://universe84a.com/collection/mucormycosis/
Mucor structures under the microscope Showing sporangium, sporangiospores, columella, sporangiophore, or aerial hyphae
Pathogenicity
Mucormycosis is a serious but rare fungal infection caused by three genera of class micromycetes, Mucor, Rhizopus, and Absidia. These fungi are saprophytes of soil, manure, and decaying vegetables. Mucormycosis mainly affects people who with weakened immune systems. It most commonly affects the sinuses or the lungs after inhaling fungal spores from the air, or the skin after the fungus enters the skin through a cut, burn, or another type of skin injury. Absidia may also cause keratitis. However, it can occur in nearly any part of the body even bone to brain. Gastrointestinal mucormycosis may occur in malnutrition, uremia, and diarrhoeal diseases.

 

 

Trichophyton mentagrophyte
Isolated: features-
The helical pattern on LPCB Mount seen
Urease test-Positive
Hair perforation test-Positive

 

 

Microsporum ferrugineum-
Growth on SDA
LPCB tease mount under microscopy

 

 

 

Epidermophyton floccosum is an anthropophilic dermatophyte (preferring humans to other hosts) that can be found worldwide.

Epidermophyton in LPCB tease mount

 

Geotricum growth on SDA and its fungal structures on LPCB preparation

LPCB Mount Footages  of Geotricum

Geotricum candidum is an extremely common fungus and worldwide in distribution. It is normal human flora and was first described in 1809 by Johann Heinrich Friedrich Link and isolated from sputum and feces. Geotricum candidum showing arthroconidium in LPCB  mount as shown above picture. Hyphae are hyaline, septate, branched, and break up into chains of hyaline, smooth, one-celled, subglobose to cylindrical arthroconidia. They are 6-12× 3-6 µm in size. They are released by the separation of a double septum.

Keynotes

  1. Geotricum is different from Trichosporon is due to lacking ability of to produce blastoconidia.
  2. Distinguishing features of G. candidum from G. capitatum is as follows-

G. candidum                                          G. capitatum

  • Risk group 1 organism                       Risk group 2
  • Cylindrical arthroconidia                  Rectangular  arthroconidia
  •  Ferment trehalose                               Can not ferment trehalose
  •  It does not assimilate D-xylose.    It assimilates D-xylose.

 

Fungus Malassezia on SDA, saline, and LPCB mount showing yeast cells and hyphae

LPCB Mount Footages of Malassezia

Malassezia growth on SDA as shown above picture. Malassezia is a monophyletic genus of fungi known to man for more than 150 years as a commensal and pathogen. French scientist Louis-Charles Malassez identified Malassezia in the late 19th century. Raymond Sabouraud identified a dandruff-causing organism in 1904 and called it “Pityrosporum malasseez”, honoring Malassez. They are a ubiquitous component of the human skin microbiome and are associated with a myriad of skin problems, including dandruff in billions of people and rarely found in places other than on the skin. 7 billion people are involved with a variety of conditions, including dandruff, atopic eczema (AE) /dermatitis, pityriasis versicolor, seborrheic dermatitis (SD), and folliculitis. In immunocompromised hosts they can also cause systemic infections.

Keynotes

  1. Among the Basidiomycota, only Malassezia and Cryptococcus are frequent human pathogens.
  2. Earlier, the hyphae filaments used to be called ‘Malassezia’, and the yeast forms were called ‘Pityrosporum’, but mycologists eventually realized they were the same organism.
  3. Catheter-related bloodstream infections (CRBSI) due to Malassezia among newborns receiving total parenteral nutrition (TPN), and in immunocompromised elderly receiving parenteral lipid supplements has also been reported.
  4. Fungemia and sepsis due to Malassezia furfur and M. pachydermatis may occur particularly in the immunocompromised elderly.
  5. M. pachydermatis, which is lipophilic but not lipid-dependent.
  6. Molecular-based methods such as Polymerase Chain Reaction techniques, and Matrix-Assisted Laser Desorption/Ionization—Time Of Flight mass spectrometry, and the chemical imprint method Raman spectroscopy is very useful tools for species identification.
  7. Molecular Identification: ITS and D1/D2 sequencing may be used for accurate species identification.
  8. MALDI-TOF MS: Capable of identifying all 14 Malassezia species in concordance with those of ITS sequence analyses.
  9. Malassezia species are difficult to grow in the laboratory on ordinary mediums lacking natural or olive oil so scrapings may be reported as ‘culture-negative since they grow best if a lipid such as olive oil is added to Littman agar culture medium or SDA.

 

 

Bipolaris growth on SDA and its structures on LPCB preparation

Bipolaris on SDA  showing colonies are moderately fast-growing, effuse, grey to blackish brown, suede-like to floccose with a black reverse. It is a large genus of dematiaceous hyphomycetes with more than 100 species, most of them being saprobes in soil and pathogens of plants, while some of the saprobic species are potentially able to infect humans and animals.

Microscopic morphology shows sympodial development of hyaline to deep olivaceous pigmented, pseudoseptate conidia on a geniculate or zig-zag rachis. Conidia mostly curved, canoe-shaped, fusoid or obclavate, rarely straight, 2–14 pseudoseptate (usually more than 6), germinating only from the ends (bipolar).

Clinical Features of Bipolaris Infection

They are allergic sinusitis, keratitis, endophthalmitis, onychomycosis, peritoneal dialysis-associated peritonitis, lung and skin infections, and, less frequently, central nervous system (CNS) infections.

Laboratory Diagnosis 

Specimen: It depends on the site of infection. e.g. nasal swab in case of allergic sinusitis whereas eye swab in case of keratitis and skin scrapping in case of skin infections.

KOH mount: For observation of fungal elements

Fungal culture: SDA or PDA can be used for fungal culture.

LPCB Preparation:  LPCB stain stands for lactophenol cotton blue and it is a combination of fixative, staining, and clearing agent. LPCB uses both as a mounting fluid and a stain. This is used for staining and microscopic identification of fungi. Its contents functions are as follows- Lactic acid: It helps in preserving the morphology of the fungal elements. Phenol: It acts as a disinfectant. Cotton blue: It stains the fungal elements as well as intestinal parasitic (cyst, ova, and oocyst) and non-parasitic structures (vegetable cells, mucus, muscle fibers, and other artifacts). Glycerol: It is a hygroscopic agent that prevents drying.

Molecular Identification: Internal transcribed spacer (ITS) region of the nuclear ribosomal DNA (rDNA) sequencing may be used to identify clinical species. The glycerol-3-phosphate dehydrogenase (GPDH) gene family has been determined to be the best single phylogenetic marker of Bipolaris species.

Keynotes on Bipolaris

  1. Key Features of Bipolaris: Dematiaceous hyphomycete producing sympodial, pseudoseptate, pale brown, straight, fusiform to ellipsoidal conidia, which are rounded at both ends.
  2. The typical morphological features of Bipolaris species include rapidly growing dark colonies, geniculate conidiophores with sympodial conidiogenesis, and large conidia with transverse distosepta, usually without a protuberant hilum and with bipolar germination. Morphologically similar anamorphic genera are Drechslera, Curvularia, and Exserohilum.
  3. Morphological differentiation of the genera relied upon a combination of characters including conidial shape, the presence or absence of a protruding hilum, the contour of the basal portion of the conidium and its hilum, the point at which the germ tube originates from the basal cell, and, to a lesser degree, the sequence and location of the first three conidial septa.
  4. There is no clear morphological boundary between genera Bipolaris and Curvularia and some species show intermediate morphology and thus using a combined internal transcribed spacer (ITS) region of the nuclear ribosomal DNA (rDNA)  and GPDH gene analysis for definitive identification of species.

 

Cladophialophora growth and its LPCB stain Microscopy

LPCB Mount Footages of Cladophialophora

Cladosporium on SDA: A Dematiaceous Fungus

Cladosporium colonies on SDA as shown above picture as shown above image. Cladosporium species colonies on Sabouraud Dextrose Agar (SDA)  and conidiophores, conidia, and microcyclic conidiogenesis with a secondary ramoconidium forming a conidiophore with a conidium attached in LPCB mount as shown in the video.

Keynote- The conidia closest to the conidiophore, and where the chain’s branch, are usually ‘shield-shaped’. The presence of shield-shaped conidia, a distinct hilum, and chains of conidia that are readily disarticulated, are characteristic of this genus as shown in the video clip.

Keynotes

  • Other genera similar to this genus are Toxicocladosporium, Penidiella, and Cladophialophora.
  • Cladophialophora species ( C. bantiana, C. carrionii and C. devriesii) are thermotolerant with lacking conidiophores and unpigmented conidial scars.
  • Cladosporium species grow easily on medium containing 10% glucose or 12–17% NaCl because of being osmotolerant.
  • Odor in these organisms is due to volatile organic compounds (VOCs) and they do not produce any major mycotoxins.

 

Key features of Curvularia spp.
Dematiaceous hyphomycete producing sympodial, pale brown, cylindrical, or slightly curved phragmoconidia, with one of the central cells

Curvularia lunata is even being a fungal plant pathogen that can cause disease in humans and other animals. The anamorph of this fungus is Curvularia lunata, while Cochliobolus lunatus denotes the teleomorph or sexual state. They are, however, the same biological entity. C. lunata is the most commonly reported species in clinical cases of reported this infection. The genus Curvularia contains some 35 species which are mostly subtropical and tropical plant parasites; however, three ubiquitous species, C. lunataC. pallescens, and C. geniculate have been recovered from human infections.

 

 

Fusarium growth on SDA and its structures in LPCB preparation

Fusarium microconidia and conidiophores in LPCB as shown above picture. Colonies are usually fast-growing, pale, or bright-colored (depending on the species) with or without a cottony aerial mycelium. The color of the thallus varies from whitish to yellow, pink, red, or purple shades. Species of Fusarium typically produce both macro-and microconidia from slender phialides. Macroconidia are hyaline, two to several-celled, fusiform to sickle-shaped, mostly with an elongated apical cell and pedicellate basal cell. Microconidia are one or two-celled, hyaline, smaller than macroconidia, pyriform, fusiform to ovoid, straight or curved. Chlamydospores may be present or absent.

 

Fusarium solani in LPCB mount Showing Micronidia on long Phialides, Macronidia, and Chlamydospores

LPCB Mount Footages of Fusarium solani

 

 

 

 

 

Penicillium in LPCB Mount Microscopy showing Condia, Conidiophores, Phialides, metulae, Septate hyphae
Penicillium structures at 400X, 800X and 1600X

LPCB Mount Footages of Penicillium

Penicillium species colony characteristics on SDA as shown above picture and colonies are usually fast-growing, in shades of green, sometimes white, mostly consisting of a dense felt of conidiophores. They are cosmopolitan in distribution and are commonly called green mold. They are found in a variety of habitats like citrus fruits, jellies, foot stuff, old leather, paper, soil, etc. Conidia of Penicillium are present everywhere in air and soil and Penicillium was accidentally discovered by Alexander Fleming, who was working on bacteria and his culture got contaminated by Penicillium notatum. They are risk group one organism.

 

 

Rhizopus in LPCB Mount with Its Various Structures

LPCB Mount Footages

Rhizopus in LPCB mount shows the presence of stolons and pigmented rhizoids, the formation of sporangiophores, singly or in groups from nodes directly above the rhizoids, and apophysate, columellate, multi spored, generally globose sporangia. After releasing of spore the apophyses and columella often collapse to form an umbrella-like structure. Sporangiospores are globose to ovoid, one-celled, hyaline to brown, and striate in many species as shown in the video.

 

 

 

Conidiophores and conidia of Scedosporium in LPCB Preparation

LPCB Mount Footages of Scedosporium

 

 

 

 

Sporothrix schenckii under the microscope
LPCB preparation
showing following structures-
conidia, conidiophores and septate hyphae
Conidia in clusters

Sporothrix schenckii is a thermodimorphic fungus widely is widely distributed in soil and in living as well as dead decaying environments. It is a causative agent of sporotrichosis, in both animals and humans. Sporotrichosis is a subacute or chronic infection and a cosmopolitan disease, occurring preferably in tropical and subtropical regions. Infection may also be associated with ulcerative, and suppurative lesions with skin and lymphatic system forming nodular lesions. Sporotrichosis was described for the first time by Benjamin Schenck, who was a medical student at Johns Hopkins Hospital in Baltimore in 1898. After the etiological isolation, Schenck sent the sample to the mycologist Erwin Smith who concluded that the agent was a microorganism of the genus Sporotrichum. The disease was reported for the second time in 1900 by Hektoen and Perkins, who classified the etiological agent as S. schenckii, with the pathogen isolated from the specimen suctioned from the skin lesions of the patient (Hektoen and Perkins 1900). The disease is also known as Rose Gardener’s disease.

 

 

 

Arthroconidia of Trichosporon inkin – Long Cylindrical in Shape

Introduction of Trichosporon

The fungus Trichosporon (genus) is characterized by the development of structures like hyaline, septate hyphae that fragment into oval or rectangular arthroconidia. The colony characteristics are suggestive for the usually raised colony and have a waxy appearance, which develops radial furrows and irregular folds.

Pathogenicity of Trichosporon

6 species of medical importance are listed below-

  1. Trichosporon asahii
  2. Trichosporon asteroides
  3. Trichosporon cutaneum
  4. Trichosporon inkin
  5. T. mucoides and
  6. T.  ovoides.

They are a minor component of normal skin flora and are widely distributed in nature. They are regularly associated with the soft nodules of white piedra and have been involved in a variety of opportunistic infections in immunosuppressed patients.

Disseminated infections are most frequently  caused by T. asahii and have been associated  with the following conditions like leukemia,

organ transplantation

multiple myeloma

aplastic anemia

lymphoma

solid tumors and

AIDS.

Disseminated infections are often fulminated and widespread, with lesions occurring in the liver, spleen, lung, and gastrointestinal tract. Infections in non-immunosuppressed patients include endophthalmitis after surgical extraction of cataracts, endocarditis usually following insertions of prosthetic cardiac valves, peritonitis in patients on continuous ambulatory peritoneal dialysis  (CAPD), and intravenous drug abuse.

 

 

Syncephalastrum in LPCB preparation under the Microscope

The fungus Syncephalastrum (genus)  is available mainly from soil and dung in tropical and subtropical regions. It can also be a difficult laboratory contaminant.

Macroscopic Features of Syncephalastrum

  • Colonies are very fast-growing, cottony to fluffy, white to light grey, becoming dark grey with the development of sporangia.
  • Microscopic Features of SyncephalastrumBroad (4-8 µm in diameter), non-septate or sparsely septate hyphae, sporangiophores, merosporangia , sporangiospores (merospores), and rhizoids are visualized. Septation of the hyphae is mostly observed as the culture gets old. Sporangiophores are frequently branched and rather short. They end up in a vesicle (80 µm in diameter). Around this vesicle are the merosporangia (4-6 x 9-60 µm), which are filled with linear chains of sporangiospores. Each merosporangium contains a single row of 3-18 merospores. Merospores (3-7 µm, may rarely reach 10 µm in diameter) are one-celled and spherical to cylindrical in shape.  Zygospores, when produced, are black, spherical, and 50-90 µm in diameter. They have conical projections. In brief, zygomycete producing sympodially branching sporangiosphores with terminal vesicles bearing merosporangia. Under the LPCB mount, here, we can see  Syncephalastrum racemosum having features like terminal vesicles, merosporangia, and merospores as shown above picture.
  • Note: The sporangiophore and merosporangia of Syncephalastrum species may confuse for an Aspergillus species when the isolate is not looking at carefully.
  • Pathogenicity and Clinical Significance
  • Syncephalastrum is a very rare causative agent of human zygomycosis. It has so far been isolated in cases of cutaneous infection and otomycosis. It comes under risk group-2 organism.

Laboratory Diagnosis of  Syncephalastrum

Specimen

  • Skin scrapping
  • Ear discharge

Direct microscopy

  • KOH preparation

Culture

  • Growth on SDA is very fast-growing, cottony to fluffy, white to light grey, becoming dark grey with the development of sporangia.

LPCB preparation 

For observation of fungal structures from culture.

 

 

Fungus, Acremonium on SDA and LPCB preparation
Colony morphology of Acremonium
Fungal structures of Acremonium under the microscope LPCB preparation

Morphology
Acremonium spp. is a slow grower and moderate growth on Sabouraud agar without cycloheximide. The colonies are white-gray or rose in color, with a velvety to cottony surface. The conidia may be single-celled ( ameroconidia), in chains, or in conidial masses, arising from short, unbranched, single, tapered phialides.

Laboratory Diagnosis 

Laboratory procedures in diagnostic mycology are directed mainly towards:

  • Microscopy: Direct demonstration of the pathogenic fungi in clinical
    specimens e.g.  KOH mount
  • Culture: Successful isolation of pathogenic fungi and fungal elements observation through LPCB preparation
  • Serology and molecular test: Supportive evidence of specific fungal infection (antigen, antibody, cell wall markers such as ß 1-3 Glucan and metabolite detection). However, PCR for the detection of fungal DNA is yet to be validated for routine diagnosis of any fungal infection.
  • Antifungal susceptibility (AFST): Prediction of possible therapeutic outcome
  • Epidemiology: Tracing the source of infection

 

 

Morphology of Ochroconis

Macroscopic characteristics on SDA: Colonies restricted, velvety to funiculose, brown to olivaceous, often with rust-brown reverse. Microscopic properties under the microscope ( LPCB preparation ): Hyphae smooth- or somewhat rough-walled, pale olivaceous. Conidiophores slightly or conspicuously differentiated, cylindrical, often flexuose, producing conidia on scattered, cylindrical to conical denticles. After detachment, an inconspicuous frill often remains both on the denticle and on the conidium base. Conidia one to four-celled, pale olivaceous brown, smooth- or rough-walled, ellipsoidal, cylindrical, clavate, or cuneiform.

Pathogenicity 

O. gallopava infection has been very rare, observed in immunologically normal people. Unlike infections in immunocompromised individuals and thus it has been increasingly recognized as a pathogen for humans following solid organ transplantation, such as kidney, liver, heart, and lung. It remains an extremely uncommon agent of human disease. When it does occur in humans, a wide range of sites may become involved, including the lung, heart, brain, superficial cutaneous or subcutaneous areas, and other parts of the body. Like other melanized fungi, the clinical presentation of O. gallopava is phaeohyphomycosis characterized by darkly colored lesions in affected tissues.

 

 

 

The fungi, Trichoderma species are common plant saprophytes and also wood-decaying fungi. Allergic presentations associated with Trichoderma exposure have been reported elsewhere. However, Trichoderma also appears to belong to the growing list of emergent etiological agents, with an increasing number of reports of invasive infections. These infections are distinguished by the presence of fine septate hyphae in tissue sections, the so-called “hyalohyphomycosis pathological entity,” for which differential diagnosis with invasive aspergillosis is condemnatory. LPCB tease mount of culture from a clinical specimen, sputum of COPD patient has shown this below video.

Conidia, Phialides and Conidiophores of Trichoderma in LPCB Preparation

 

 

 

Contd…LPCB Mount Footages of Various Fungi

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