MLS 314 (MYCOLOGY AND VIROLOGY) – WEEK 1 & WEEK 2 Flashcards
1
Q
the asexual form of a fungus
A
Anamorph:
2
Q
a specialized conidiogenous cell from which a succession of spores is produced and which has a column of apical scars at its tip.
A
• Annellide
3
Q
: ability of a fungus to use a specific carbon or nitrogen source for growth.
A
•Assimilation
4
Q
: without cross-walls or septa.
A
•Aseptate
5
Q
: the process of conidium formation.
A
•Conidiogenesis
6
Q
[in which an existing hyphal cell is converted into one or more conidia]
A
thallic
7
Q
[in which conidia are produced as a result of some form of budding process]
A
blastic
8
Q
An asexual (mitosis only) propagule that forms on the side or the end of the hypha or conidiophore.
A
Conidium
9
Q
It may consist of one or more cells.
A
Conidium
10
Q
It is always borne externally, ie., not enclosed in a saclike structure such as sporangium.
A
Conidium
11
Q
If a fungus produces two types of conidia, those that are small and usually singled cell are referred to as (?)
A
microconidia
12
Q
are usually segmented into two or more cells
A
macroconidia
13
Q
: darkly pigmented.
A
•Dematiaceous
14
Q
: ability of a fungus to utilize a specific carbohydrate in the presence of other organic compounds, resulting in the production of gas.
A
•Fermentation
15
Q
(color change) may simply indicate that the carbohydrate has been assimilated.
A
Acid production
16
Q
All carbohydrates (?) by a fungus are also assimilated, but many compounds that are (?) are not necessarily fermented.
A
fermented
assimilated
17
Q
: colonies with a cotton-like texture.
A
•Floccose
18
Q
: term used to describe spores with a spindle-like shape.
A
•Fusiform
19
Q
: colonies with wax-like texture.
A
•Glabrous
20
Q
: a self-sterile fungus; sexual reproduction cannot take place unless two compatible mating strains are present.
A
•Heterothallic
21
Q
: a self-compatible fungus; sexual reproduction can take place within an individual strain.
A
•Homothallic
22
Q
: colorless, transparent, transluscent.
A
•Hyaline
23
Q
: one of the individual filaments that make up the mycelium of a fungus.
A
•Hypha (pl., hyphae)
24
Q
: a filamentous fungus.
A
•Mold
25
: hyaline or lightly colored.
•Moniliaceous
26
: a mass of branching filaments which make up the vegetative growth of a fungus.
•Mycelium
27
: a cell with several nuclei.
•Oligokaryotic
28
: having few septa.
•Pauciseptate
29
: a chain of yeast cells which have arisen as a result of budding and have elongated without becoming detached from each other, forming a hypha-like filament. No marked constriction.
•Pseudohypha (pl., pseudohyphae)
30
: a short branching hypha that resembles a root.
•Rhizoid
31
: having cross-walls or septa.
•Septate
32
: a monophyletic clade of species with equivalent clinical relevance.
•Species complex
33
: a specialized hypha upon which a sporangium develops.
•Sporangiophore
34
: term used to describe the development of a single conidium at successive sites along a lengthening conidiogenous cell.
•Sympodial
35
: the sexual form of a fungus.
•Teleomorph
36
: one of the two basic forms of conidiogenesis.
•Thallic
37
: the vegetative growth of a fungus.
•Thallus
38
: a unicellular, budding fungus.
•Yeast
39
named species
undescribed species
new species described each year
135,000
1 million to more than 10 million
1,000 to 1,500
40
human and zoonotic diseases
opportunistic infections
< 500
< 50
41
Importance of
1.Increasing number of ubiquitous environmental molds implicated as (?) capable of producing serious diseases among debilitated or immunocompromised hosts.
2.One of the leading causes of (?)
3.Often mistaken for as (?) that produces fatal consequences.
4.Increase (?) (travel to an endemic area or a fungus that exists as part of indigenous flora of the local population)
5. (?) population.
opportunistic pathogens
nosocomial infections
bacterial infection
morbidity
Aging
42
: Fungi were used as an antiseptic and anesthesia due to the “magical & spiritual” properties.
•35,000 BC
43
: People were convinced that association with fungi will entail the formation of diseases. People believed that fungi were “the work of the devil”.
•Middle Ages
44
: Fungi were plants with no fruit nor seed.
•Renaissance period
45
: Birth of the 1st mycological studies. Pier Anton Micheli— founder of modern mycological studies. Mycology was separated from botany.
•18th Century
46
: Fungi were recognized as a potential causative agents of diseases that are usually fatal in nature.
•Mid-20th Century
47
: branch of science that deals with systemic biological classification of all living organisms and divided into three disciplines viz., classification, nomenclature, and identification.
•Taxonomy (Gk.: taxis = arrangement; anomia = method)
48
•Taxonomy (Gk.: (?) = arrangement; (?) = method)
taxis
anomia
49
•The scientific names of fungi are subject to the (?) that was adopted in 2011.
International Code of Nomenclature (ICN)
50
•In 2011, a breakthrough in the development of (?) concept recommended the discontinuation of the dual nomenclature system for fungi with anamorphic and teleomorphic forms.
“One Fungus, One Name”
51
Effective January 1, 2013 the system of permitting separate anamorph and teleomorph names ended; and assigned priority to the (?) independently of whether the organism was originally described as the anamorph or the teleomorph.
oldest genus (species) name
52
•Eukaryotes are divided into five monophyletic lineages or supergroups as proposed by Adl et.al.1
1.SAR (one clade consists of three groups viz., Stramenopiles, Alveolata, and Rhizaria)
2.Archaeplastida
3.Excavata
4.Amoebozoa
5.Opisthokonta — True fungi
53
— organisms that are not fungi sensu strictu (eg., Rhinosporidium seeberi) that share fungal-like morphological features with the true fungi.
Parafungi or pseudofungi
54
Fungi Imperfecti also termed (?)— fungi that reproduced asexual type only (sporogenesis), while perfect fungi are capable of both sexual (teleomorph) and asexual (anamorph) type of reproduction.
“form-division Deuteromycota”
55
Basic scheme of hierarchical organization of the Kingdom Fungi
Eukarya
Fungi (Mycetea)
- mycota (Basidiomycota), - mycotina
- mycetes (Agaricomycetes)
- ales (Agaricales)
- ace (Agaricaceae)
- Agaricus
- Agaricus bisporus
56
Seven phyla that constitute the true fungi
1.Ascomycota
2.Basidiomycota
3.Blastocladiomycota
4.Chytridiomycota
5.Glomeromycota (formerly Zygomycota)
6.Microsporidia
7.Neocallimastigomycota
57
•Consists of four subphyla incertae sedis.
Glomeromycota (formerly Zygomycota)
58
accommodates Mucorales that includes the genera Lichtheimia (formerly Absidia), Mucor, Rhizomucor, and Rhizopus.
Subphylum Mucormycotina
59
has been created for the Entomophthorales that includes the genera Basidiobolus and Conidiobolus — agents of subcutaneous infections
Subphylum Entomophthoromycotina
60
Most members are septate, filamentous thallus, but some are atypical yeasts.
Phylum Basidiomycota
61
produced via sexual reproduction of a generative cell (basidium).
Basidiospores (haploid)
62
The most prominent are the basidiomycetous yeast with anamorphic stages belonging to the genera:
1.Cryptococcus
2.Malassezia
3.Trichosporon
63
Polyphyletic, belong to subphylum Agaricomycotina, class Tremellomycetes, and order Tremellales and(or) Filobasidiales1
Cryptococcus
64
Asexual reproduction is variable; do not produce spores like those of the Ascomytoca.
Phylum Basidiomycota
65
Most filamentous basidiomycetes are wood-rotting fungi or obligate plant pathogens.
Phylum Basidiomycota
66
Contains ~50% of all named fungal species and accounts for ~80% of fungi of medical importance.
Phylum Ascomycota
67
Sexual reproduction leads to the development of haploid spores, termed (?) which are produced in a sac like structure called (?). (?) contains numerous asci.
ascospores
ascus
Ascocarps or ascomata
68
Asexual reproduction leads to conidiation from a generative conidiogenous cell.
Phylum Ascomycota
69
Three medically important subphylum:
1.Taphrinomycotina
2.Saccharomycotina
3.Pezizomycotina
70
Taphrinomycotina contains the genus (?) formerly classified under kingdom Protozoa.
Pneumocystis
71
Saccharomycotina contains the class (?) belongs to order Saccharomycetales
Saccharomycetes
72
Pezizomycotina contains two classes: (1) and the (2).
1. Eurotiomycetes
2. Sordariomycetes
73
Class Saccharomycetes
Phylum Ascomytoca
74
Characterized by vegetative yeast cells which proliferate by budding or fission.
Class Saccharomycetes
75
Do not produce ascomata, the ascus being formed by direct transformation of a budding vegetative cell, by “mother-bud” conjugation, or by conjugation between two independent single cells.
Class Saccharomycetes
76
Members include the anamorphic stage belonging to genus Candida containing ~200 anamorphic species and has teleomorphs in more than 10 genera.
Class Saccharomycetes
77
Belonging to class Eurotiomycetes or Sordariomycetes Teleomorphs of melanized fungi:
4. Capnodiales
5. Chaetothyriales
6. Microascales
7. Pleosporales
8. Ophiostomatales
78
basic structural unit consists of a chain of multinucleate, tubular, filament-like cells.
Hypha
79
Most multicellular fungi in their vegetative state, consists of a mass of branching hyphae called (?).
mycelium or thallus
80
Each hypha has a rigid cell wall and increases in length as a result of apical growth with mitotic cell division.
Multicellular fungi
81
In a more primitive fungi, the hyphae remain aseptate (coenocytic).
Multicellular fungi
82
In a more advanced groups, the hyphae are divided into compartments or cells by the development of more or less frequent cross-walls, termed septa.— septate hyphae.
Multicellular fungi
83
Fungi that exists in the form of microscopic multicellular mycelium are commonly referred to as (?).
molds
84
Exists in the form of independent single cells propagate by budding out similar cells from their surface.
Unicellular fungi
85
The bud may become detached from the parent cell or it may remain attached and itself produce another bud, leading to the production of a chain of cells or described as loosely arrangement of budding cells —
yeasts
86
Under certain conditions, continued elongation of the parent cell before it buds out results in a chain of elongated cells, termed (?).
pseudohypha
87
Compare to true hypha, the connection between adjacent they shows a marked constriction.
pseudohyphal cells
88
Medically important fungi that change their growth form as a part of the process of tissue invasion.
Dimorphic fungi
89
Dimorphic pathogens change from a multicellular mold form (25-30C) in the natural environment to a budding, single-celled yeasts form (35-37C) in tissue under the influence of temperature— (?).
thermal dimorphism
90
Dimorphic fungi
1.Histoplasma capsulatum
2.Blastomyces dermatitidis
3.Coccidioides immitis
4.Paracoccidioides brasiliensis
5.Sporothrix schenckii
91
Cell wall
Rigid, mostly composed of 90% polysaccharides eg., (?); 10% (?) in various combinations.
chitin, glucan, chitosan, galactosan, and mannan
proteins and glycoproteins
92
the major carbohydrate consist of repeating monomers of N-acetyl-glucosamine (NAG) which provides shape and protection from osmotic lysis (unaffected by some antibiotics).
Chitin
93
Barrier between fungal cell and its external environment.
Cell wall
94
Binding site for some enzymes
Cell wall
95
Possesses antigenic properties which allow interaction with other organisms
Cell wall
96
Composed of structurally arranged phospholipids in two-layered configuration scattered randomly.
Cell membrane
97
major component that regulates solute intake and secretion (transport system) through selective permeability and serves as the target of antifungal drugs like Nystatin.
Ergosterol
98
Facilitates biosynthesis of cell wall and capsular material.
Cell membrane
99
Protects the cytoplasm
Cell membrane
100
External coating located outside or covering the cell wall and found only in certain fungi.
Capsule
101
Composed of amorphous polysaccharide.
Capsule
102
May influence growth of fungus by preventing dissociation of buds and dispersion of yeast.
Capsule
103
Virulence factor
Capsule
104
Contains the following organelles:
1.Nucleus
2.Chromosomes
3.Ribosomes
4.Mitochondrion
5.Endoplasmic reticulum
6.Vacuoles
Cytoplasm
105
bounded by a nuclear membrane, may vary in size, shape, and number. Usually contains one nucleolus of mostly RNA.
Nucleus
106
described as linear.
Chromosomes
107
present at all times for protein synthesis.
Ribosomes
108
generation of energy.
Mitochondrion
109
Require preformed organic carbon compounds for nutrition.
Nutritional type is chemoheterotrophic (lacks chlorophyll)
110
Obtain their nourishment by secreting enzymes into the external substrate and by absorbing the released nutrients through their cell wall.
Nutritional type is chemoheterotrophic (lacks chlorophyll)
111
Capable of growing on dead (saprophytic) and living organic matter.
Nutritional type is chemoheterotrophic (lacks chlorophyll)
112
Usually nonmotile except for the phylum Chytridiomycota and species of Rhizophidium.
Motility and atmospheric requirements
113
Mostly aerobic, some are facultative anaerobic.
Motility and atmospheric requirements
114
Lysine synthesis
Utilize -aminodiphate pathway.
115
Lysine synthesis
Precursor for (?), also for the development of (?).
penicillin and lysine
antifungal drugs
116
describes the propagules that result form an asexual process (mitosis only) and generally short-lived propagules.
Conidium (pl. conidia)
117
Most fungi are capable of [?] reproduction (involving meiosis, preceded by fusion of the nuclei of two cells).
sexual
118
Some species are [?] (self-fertile) and able to form sexual structures between different cells within an individual thallus.
homothallic
119
Most are (?) that do not form their sexual structures unless two capable isolates come into contact
heterothallic
120
— once two compatible nuclei have fused, meiosis can occur leading to the production of sexual (?).
Spores
121
In some cases, spores are produced in millions in macroscopic “fruiting bodies” such as (?).
mushrooms
122
Cell wall of Fungi
Polysaccharides (ie., Chitin, Glucan, Mannan, Galactosan, Chitosan)
123
Cell wall of Bacteria
Polysaccharide
Peptidoglycan
124
Cell membrane of Fungi
Ergosterol
125
Cell membrane of Bacteria
No sterols, EXCEPT: Mycoplasma and Ureaplasma
126
Nucleus of Fungi
Small, bound by nuclear membrane
127
Nucleus of Bacteria
Nucleoid, no nuclear membrane
128
Chromosome of Fungi
Linear
129
Chromosome of Bacteria
Circular
130
Ribosomes of Fungi
80s
131
Ribosomes of Bacteria
70s
132
Lysine synthesis of Fungi
-aminodiphate pathway
133
Lysine synthesis of Bacteria
DAP (Diaminopimelate pathway)
134
Identification of Yeasts
1.Morphological
2.Physiological, and
3.Biochemical characteristics
135
Morphological includes:
a.color of colonies
b.size and shape of cells
c.presence of capsule around the cell
d.production of hyphae, pseudohyphae, arthroconidia, and(or) chlamydospores
136
Biochemical characteristics
a.assimilation and fermentation of sugars
b.assimilation of nitrate
137
•Most commercial tests (eg., ID YST, bioMérieux) are based on sugar assimilation of isolates.
•DNA sequencing
•Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF)
Identification of Yeasts
138
Identification of Molds
Macroscopic characteristics viz. colonial form, surface color (obverse) pigmentation, reverse pigmentation, and growth rate.
139
Factors that influence the colonial form:
1.Culture medium
2.Incubation temperature
3.Age of culture
4.Amount of inoculum
140
is important in speciation, hence it is important to select culture conditions which favor sporulation.
Sporulation
141
promotes over production of mycelium which results in loss of sporulation, in such cases, subculture to a low-nutrient medium (stimulate sporulation)
Sabouraud’s dextrose agar (SDA)
142
Compose of the basic unit of fungus (hyphae).
Mold cells
143
The septum, represents the cross walls (divisions) of a hyphae.
Mold cells
144
Mass of hyphae will produce mycelium.
Mold cells
145
Cells are mostly spherical to ellipsoidal of 3 to 15 microns in size.
Yeast cells
146
Multiply via asexual reproduction known as (?).
budding
147
Stages of budding:
1.Bud emergence >
2.Protuberance
3.Elongation.
148
Growth can be observe through obverse (surface) or reverse (back). Obverse observation can be white to cream, bright or light gray to brown, reverse may be non pigmented, to yellow, orange, or red.
Mold cells
149
•As seen in culture, colonies may appear as white opaque, pasty to creamy to mucoid (encapsulated) with 0.5 to 3mm in diameter. •Colonies grow at 35-37C, optimum
Yeast cells
150
are fungus with cross walls
Septate hyphae
151
are nearly parallel to one another with dichotomous branching at acute angles — Aspergillus spp.
Septate hyphae
152
are fungus without cross walls. Coenocytic hyphae results form nuclear division within a cell without division in the cytoplasm.
Aseptate
153
Multiple projections in a hypha resembling an old comb hyphae appearance
Pectinate
154
Hypha with club-shaped cells, the layer end of one cell being attached to the smaller end of an adjacent
Racquet
155
•Hypha forming coiled or corkscrew like turns.
•Characteristics of Trichophyton mentagrophytes
Spiral
156
Terminal hyphae branches that are irregular, broad, and antler-like in appearance. Especially characteristics of T. schöenleinii.
Favic chandelier
157
Round knot-like structure formed by intertwined hyphae; seen especially among dermatophytes
Nodular
158
•Root like structure along the vegetative hyphae
•Characteristics of certain Glomeromycota (formerly Zygomycota) (Rhizopus and Absidium)
Rhizoids
159
Clear, transparent, and colorless hyphae
Hyaline
160
Hyphae having structures that are brown to black, this is due to the melanotic pigment in the cell walls
Dematiaceous
161
Triggered by stems from the sub apical accumulation of wall precursors (presumable vesicles) reaching a critical concentrations.
Lateral Elongation
162
i.Cell vesicles move towards the tip of the hyphae (towards the plasma membrane) where they release various enzymes and other compounds.
ii.Enzymes release by these particles involved in the lysis and synthesis of the cell wall
iii.Following lysis of the cell wall, cell division and growth allows for the hyphae to be elongated at the tip.
iv.Once the new cell has formed, enzymes at the tip start synthesizing a new cell wall around the new cell to protect it. Followed by strengthening of the actin cap. Then disappearance of Spitzenkörper.
v.Spitzenkörper, play an important role in organizing (regulating) hyphal growth. Found behind hyphal tip (apex).
Process of apical elongation
163
(?) move towards the tip of the hyphae (towards the plasma membrane) where they release various enzymes and other compounds.
Cell vesicles
164
(?) release by these particles involved in the lysis and synthesis of the cell wall
Enzymes
165
Following lysis of the (?), cell division and growth allows for the hyphae to be elongated at the tip.
cell wall
166
Once the new cell has formed, enzymes at the tip start synthesizing a new (?) around the new cell to protect it. Followed by strengthening of the actin cap. Then disappearance of Spitzenkörper.
cell wall
167
(?), play an important role in organizing (regulating) hyphal growth. Found behind hyphal tip (apex).
Spitzenkörper
168
leads to the formation of filamentous (fuzzy) colonies — mycelium
Apical elongation
169
Types of mycelium:
1.Vegetative
2.Aerial mycelium
170
filamentous (fuzzy) colonies
mycelium
171
buried down, for the water exchange and nutrient absorption.
Vegetative
172
projects on the surface of the culture medium with reproductive structures known as spores which can be sexual or asexual.
Aerial mycelium
173
Requires the formation of special clusters for fertilization and nuclear fission.
Sexual reproduction
174
sex organ, gametes (sex cells) and either monoecious or dioecious
Gametangium
175
Involves meiosis
Sexual reproduction
176
are developed as a result of a primary nuclear fusion with reduction in chromosome number during their formation.
Sexual spores
177
Germinates and forms hyphae and mycelium
Sexual reproduction
178
2-8 ascospores within an ascus (sac), the shape of each aids in identification of fungus.
Ascospores
179
Basidiospores
180
When two sporangiophores sexually fuse to form a large thick-walled bodies
Zygospores
181
contains zygospores along a nonseptate hyphae
Zygosporangium
182
Produced from a fusion of 2 non-identical hyphae
Oospores
183
Sporulation and spores are preferred terms used when there is a merging of nuclear material or genes combined.
Asexual reproduction
184
spore formed by budding, no fusion of nuclei takes place in the formation of spore.
Asexual spores
185
Do not form mycelium, produce pasty type of colony (yeast)
Unicellular
186
Form mycelium, produce filamentous colony (mold)
Multicellular
187
Spores contained in sporangia or sacs are produced terminally (at the end)
Sporangiospores
188
specialized stalk which bears the sporangia
Sporangiophore
189
Observed among, Zygomycetes, Rhizopus
Sporangiospores
190
Results from the transformation of a vegetative yeast or specialized hypha called conidiophores. Often referred to as macro- (multicellular) and microconidia (unicellular)
Mitospores (Conidiospores)
191
Freed by abstraction at the point of attachment
Mitospores (Conidiospores)
192
Observed among Ascomycetes and Deuteromycetes
Mitospores (Conidiospores)
193
produced on a thallus
Thallospores
194
Formed by budding from a pseudohypha
Chlamydospores
195
Thick-walled resting spores either within (intercallary) the segments; hyphal sides (sessile); or hyphal tip (terminal)
Chlamydospores
196
highly developed conidiophores
Phialospores
197
Vesicle, enlarged swollen cell often at the end of a conidiophore or sporangiophore and it bears conidia.
Phialospores
198
The conidiospores are formed usually at the sides (tips) of a hyphae and are usually in chains.
Phialospores
199
Outer skin layer (hair)
Do not penetrate tissues
Malassezia furfur
Superficial
200
Skin, hair, nails (keratinized area)
Do not penetrate tissues, but can cause inflammation
Trichophyton species
Microsporum species
Cutaneous
201
Connective tissues, Muscles, Bones
Penetrate tissues, transmitted via traumatic inoculation
Sporothrix schenkii
Rhinosporidium seeberi
Loboa loboi
Subcutaneous
202
All body system
Thermal (dimorphic)
Histoplasma capsulatum
Systemic
203
Any part
Infects immunocompromised host
Aspergillus fumigatus
Candida albicans
Opportunistic
204
Similar to bacteriology, the goal of mycology laboratory is to accurately (?) fungi suspected of causing infection.
isolate and identify
205
Important considerations on specimen collection, transport, and processing
1.Proper (?) (aseptic technique).
2.All specimens should be transported in (?) and processed ASAP.
3. (?) or anaerobic containers should never be used for fungi.
4. (?) is recommended for transport (some fungi can be affected by above 37C and below 10C— dermatophytes are sensitive to cold temperatures.
5.If delay is expected within 2h, specimen should be stored at (?).
6.Appropriate volume. Volume required for fungal culture is greater than in bacteriology, since fluids and respiratory secretions need to be (?) prior to plating.
collection
leakproof sterile containers
Anaerobic transport media
RT
4C
concentrated or pretreated
206
the appropriate physicians should be notified.
•For rejected or unacceptable specimen
207
requisition must accompany each specimen and must include the following demographics
1.Patient name
2.Age and sex
3.Location or address
4.Physician name
5.Specific culture site
6.Date and time of specimen collection
7.Clinical diagnosis
8.Any special culture request
208
Specimen must have a firmly attached label indicating the
patient name, specimen type, date and time of collection
209
Necessary to maximize recovery of fungi.
Pretreatment and optimum temperature for culture
210
Importance of Pretreatment and optimum temperature for culture
1.Release fungi enclosed within cells.
2.Concentrate fungal material in the specimen.
3.Help to reduce or eliminate bacteria present in contaminated specimens because of the action of mucolytic agents viz.
211
mucolytic agents
a.N-acetyl-L-cysteine
b.5% oxalic acid
c.Dithiothreitol (Sputolysin2)
212
Temperature requirements for culture:
1.Blood, CSF: 30-37C
2.Dermatological (ie., skin, hair, nails): 15-30C
213
Granules should be washed & crushed; other materials should be centrifuge at 2,000g for 10min.
Abscess, drainage, pus, granules
214
Lysis in Isolator tubes and then centrifugation for 30min. at 3,000g, in a 35 fixed-angle rotor or swing bucket.
Blood, bone marrow
215
Criteria for the detection of H. capsulatum and other dimorphic fungi.
Blood, bone marrow
216
Centrifugation at 2,000g for 10min. or membrane filtration.
Body fluids
217
Essential for best recovery with volumes 1mL; blood clots should be teased apart.
Body fluidsC
218
Mince; gently pushed pieces down into agar.
Nails
219
Essential for the recovery of dermatophytes.
Nails
220
Lysis with mucolytic agents, followed by centrifugation at 2,000g for 10min.
Respiratory secretions (BAL, sputum)
221
Critical for the recovery of Pneumocystis jirovecii as very difficult to isolate in culture; improves recovery of other mycoses.
Respiratory secretions (BAL, sputum)
222
Mince or grind in a mortar; push pieces down into agar
Tissue
223
Essential for best recovery; for zygomycetes and other molds, mincing is best; for H. capsulatum, grinding is best.
Tissue
224
Centrifugation at 2,000g for 10min.
Urine
225
Essential for best recovery, particularly deep mycoses.
Urine
226
Essential for best recovery.
Abscess, drainage, pus, granules
227
•All work in mycology should be carried out in a certified (?) especially when dealing with filamentous fungi.
type 2 laminar airflow BSC
228
can be handled on the bench in the same manner that bacterial cultures are routinely handled.
Yeasts cultures
229
are recommended for personnel working with clinical specimens that may contain dimorphic fungi.
•BSL 2 procedures
230
should be worn for processing specimens and culture.
•Appropriate PPE
231
of slanted media are safer to handle than petri plates.
•Screw-cap tubes
232
should NEVER be used if Coccidioides immitis is suspected.
•Petri plates
233
•Do not make slide cultures of isolates that may be:
1.Histoplasma capsulatum
2.Blastomyces dermatitidis
3.Coccidioides immitis
4.Coccidioides posadasii
5.Paracoccidioides brasiliensis
6.Talaromyces marneffei
7.Cladophialophora bantiana
234
Clean surface with 70% alcohol. Collect from active peripheral edge with sterile needle and syringe. If open, use swab system or aspirate.
Abscess (Drainage, exudate, pus, wound)
235
If thick, pretreatment similar to sputum specimen
Abscess (Drainage, exudate, pus, wound)
236
If 2h, RT
Abscess (Drainage, exudate, pus, wound)
237
Examine for grains or granules and note color if present
Abscess (Drainage, exudate, pus, wound)
238
Disinfect skin with iodine tincture or chlorhexidine prior to obtaining. Use max. volume of blood recommended for the system used.
Blood
239
Manual
Biphasic (Septi-Chek)
Automated (BacT/ALERT, bioMérieux)
Lysis-centrifugation
Blood
240
If 2h, RT; if longer, RT but process in 16h (lysis- centrifugation)
Blood
241
All systems for bacteria will recover all yeasts except for Malassezia spp. but will not recover molds.
Blood
242
Lysis centrifugation systems are good for recovery of molds, especially the causing endemic mycoses.
Gives high contamination and false positive rates.
Blood
243
Collect aseptically in a heparinized syringe or lysis-centrifugation tube.
Bone marrow
244
Clotted bone marrow is an unacceptable specimen.
Bone marrow
245
If 15min., RT; if longer RT
Bone marrow
246
Pediatric isolator tubes are best.
Bone marrow
247
Remove distal 3-5 cm of line tip and place in a sterile container.
Catheter tip (Intravascular)
248
Method of Maki et. al.2; used for catheter tips but not validated for fungi.
Catheter tip (Intravascular)
249
If 15min., RT; if longer,4C
Catheter tip (Intravascular)
250
Avoid media containing Cycloheximide.
Catheter tip (Intravascular)
251
Disinfect all types with 70% alcohol.
Cutaneous (Hair, skin, nails)
252
is most important, plucking is best; submit 10-12 hairs in sterile dry container or envelope.
Hair
hair root
253
scrape with dull edge of a scalpel or glass slide, or vigorously brush in a circular motion with soft-bristle brush.
skin
254
material under nail should also be scraped. submit in sterile container or envelope.
nails clip or scrape with scalpel
255
Only the leading edge of a lesion should be sampled, as centers are often nonviable.
Cutaneous (Hair, skin, nails)
256
All specimens should be pressed gently into the agar with a sterile swab, do not streak agar plates. If used, toothbrushes should be pressed gently into agar as well
Cutaneous (Hair, skin, nails)
257
If 72h., (very stable) Never refrigerate, as dermatophytes are sensitive to cold.
Cutaneous (Hair, skin, nails)
258
Select hairs with fluoresce under a Wood’s light.
Cutaneous (Hair, skin, nails)
259
can be collected with soft-bristle toothbrush.
Hair and skin
260
olive oil or a paper disc saturated with olive oil should be placed on the first quadrant of agar plate.
For pityriasis versicolor (M. furfur)
261
taken by physicians and media(slides) inoculated directly.
Corneal scraping
262
needle aspiration.
Vitreous humor
263
inoculate non-inhibitory media in X- or C-shape motion.
Corneal
264
concentrate by centrifugation, use sediment for media and smears.
Vitreous humor
265
If 15min., RT; if longer RT
Eye (corneal scraping, vitreous humor)
266
Very little material usually available. Avoid media with cycloheximide
Eye (corneal scraping, vitreous humor)
267
Collected surgically. Transport in sterile container.
Medical devices
268
Use sterile scalpel to collect (by scraping) biofilm or vegetative growth.
Medical devices
269
If 15min., RT; if longer, 4C
Medical devices
270
Avoid media containing cycloheximide. Device material recovered best by using liquid medium.
Medical devices
271
Have patient empty bladder and then massage prostate gland to yield fluid.
Prostate fluid
272
Inoculate media directly or transport in sterile wide mouth container.
Prostate fluid
273
If 15min., RT; if longer RT
Prostate fluid
274
Fluid should always be examined microscopically. The first urine following massage has a high yield. This fluid is excellent for detection of endemic mycoses.
Prostate fluid
275
Use first morning sputum collected after brushing teeth.
Collect brushing and BAL fluid surgically.
Place all samples in sterile containers.
Inoculate media containing antimicrobial agents with and without cycloheximide.
Respiratory tract, lower (sputum, bronchial aspirate, BAL fluid)
276
Viscous lower respiratory specimens should be pretreated and centrifuged to concentrate their contents.
Respiratory tract, lower (sputum, bronchial aspirate, BAL fluid)
277
If 2h., RT; if longer, 4C
Respiratory tract, lower (sputum, bronchial aspirate, BAL fluid)
278
Saliva and 24h sputum are unacceptable specimens. Methods for mycobacterial decontamination are not acceptable.
Respiratory tract, lower (sputum, bronchial aspirate, BAL fluid)
279
Swab oral lesions, avoiding tongue. Use thin wire or flexible swab or oropharynx. Collect sinus contents surgically.
Respiratory tract, upper (Oral, oropharyngeal, and sinus samples)
280
Use swab transport system for oral and oropharyngeal samples. Place sinus contents in sterile container
Respiratory tract, upper (Oral, oropharyngeal, and sinus samples)
281
Oral: if 2h, RT; if longer, RT
Sinus: if 15min. RT; if longer, RT
Respiratory tract, upper (Oral, oropharyngeal, and sinus samples)
282
Selective and chromogenic media are best for recovery of Candida spp.
Respiratory tract, upper (Oral, oropharyngeal, and sinus samples)
283
Collect as for bacteriology. Concentrate by centrifugation, and use sediment for inoculation. Clots should be grounded.
Sterile body fluids (CSF and pericardial, peritoneal, and synovial fluids)
284
Except CSF, put sterile body fluids in sterile Vacutainer tubes with heparin or lysis- centrifugation tube to prevent blood clotting.
Except for CSF, blood culture bottles can be used for recovery of yeast.
Sterile body fluids (CSF and pericardial, peritoneal, and synovial fluids)
285
If 15min., RT; if longer, RT; never refrigerate.
Sterile body fluids (CSF and pericardial, peritoneal, and synovial fluids)
286
should always be examined microscopically. With specimen volumes 2ml, fluid should be plated directly using as much fluid on each plate as possible.
Sterile fluid sediment
287
Specimen use should be discourage.
Stool
288
Collected surgically. A larger volume is needed than for Bacteriology.
Tissue
289
Use sterile container, keep moist (saline drops) to prevent drying.
Except with H. capsulatum, mincing (not grinding) is critical.
pieces should be pressed into the agar so they are partially embedded.
Grind tissue for recovery of H. capsulatum.
Tissue
290
If 15min., RT; if longer, RT
Tissue
291
recommended for invasive disease.
Examine subcutaneous portion for granules (see abscesses)
Tissue biopsy
292
First morning clean catch, suprapubic, or catheterized specimens; 24-h specimens are unacceptable.
Urine
293
Use a sterile container or urine transport system.
Concentrate specimens by centrifugation, and use sediment for inoculation.
Urine
294
If 2h, RT; if longer, 4C; urine transport systems can stay at RT for up to 72h.
Urine
295
Chromogenic media best for Candida. Use sediment for microscopic examination.
Urine
296
Collect as for bacteriology.
Vaginal
297
Swab transport system or sterile container for washings.
Vaginal
298
If 2h, RT; if longer, RT
Vaginal
299
Antibacterial media or chromogenic agars are best for recovery of Candida.
Vaginal
300
Common clinical sites for laboratory recovery of pathogenic fungi
301
General approaches for direct detection and identification of fungi
1.Culture based
2.Non-culture-based
302
2.Non-culture-based
a.Direct microscopic examination
b.Antibody and antigen detection
c.Detection of 1,3--D-glucan and other fungal metabolites
d.Mass spectrometry
e.Nucleic acid detection
303
Stains and methods used directly on unfixed tissue or samples
Potassium hydroxide (KOH) (5-30 min.)
Calcofluor white, CW (1-2 min.)
Rapid Giemsa-like (Diff-Quik) (2-3 min.)
Colloidal carbon wet mounts (India ink, nigrosin) (1 min.)
Lactophenol cotton blue (LPCB)
304
•Wet mount prepared in (?), used to distinguished fungi in thick mucoid specimens or in samples that contain keratinous material such as skin, hair, and nails.
10%
305
•Nails may require stronger concentration of up to (?) to digest the nail prior to microscopy.
25%
306
•The fungi remain unaffected while the (?) of the host cells are partially digested2.
proteinaceous components
307
•Solutions of NaOH (?) may be used as an alternative to KOH.
10% or 25% with added glycerin
308
•Visualization of fungal elements may be enhanced by the addition of (?) or glucan-binding fluorescent brighteners such as (?), which bind to chitin.
lactophenol cotton blue (LCB)
calcofluor white (CW)
309
KOH, basic procedure
1. Addition of (?)
2. Specimen covered in (?)
3. Wet house for (?)
4. Observe under the (?)
10% KOH
coverslips
5 to 30 minutes
microscope
310
•Nonspecific, nonimmunological fluorochromes (along with Uvitex 2B and Blankophor) that bind to β-1,3 and β-1,4 polysaccharides specifically cellulose and chitin of fungal cell walls and fluoresces when exposed to long-wave UV light
Calcofluor white, CW
311
•More rapid than auramine-rhodamine stain.
Calcofluor white, CW
312
•Fungal elements appear bluish white to green fluorescence against a dark background when excited with UV or blue-violet radiation
Calcofluor white, CW
313
are 5-8 m [], round, and uniform in size and exhibit a characteristic peripheral cyst wall staining with an intense “double parenthesis-like” structure in CW
P. jirovecii cyst
314
•Yeast cells are differentiated from P. jirovecii by
budding and intense staining
315
fluoresce strongly and must be differentiated from fungal hyphae.
•Cotton fibers
316
1.KOH (10%) is mixed in equal proportion with CW solution (?)
(0.1 g of CW M2R and 0.05 g of Evans blue in 100 ml H2O)
317
2.Specimen is covered with this mixture and a coverslip is applied for 5 to 10 min. (nail and tissue preparations require up to (?) incubation time).
During (?), clearing and interaction of stain with the fungal elements occur.
(?) of the slide may speed up the clearing process.
30 min.
incubation
Warming
318
CW, basic procedure
3.Examination of the preparation with a (?) containing appropriate (?) at x 100 to x 400 magnification
fluorescent microscopy
excitation and barrier filters
319
may stain poorly with CW due to pigmentation that masks the fluorescence.
Dematiaceous fungi
320
Cysts walls with internal thickenings (double comma).
P. jirovecii of BAL fluid
321
Dichotomous branching hyphae.
Aspergillus spp. of lung tissue
322
Chitinous walls of the fungus.
C. albicans from culture
323
•Also applied to other specimens, such as BAL fluid or CSF.
Rapid Giemsa-like (Diff-Quik)
324
•Yeasts and trophozoites appear as blue purple
Rapid Giemsa-like (Diff-Quik)
325
(?)
•Differentiate form Leishmania;
Leishmania has a (?) and Histoplasma does not.
Rapid Giemsa-like (Diff-Quik)
Kinetoplast
326
A.Thick cluster of mostly trophic forms (2-3m) with small reddish purple nuclei and light blue to red violet cytoplasm.
Rapid Giemsa-like (Diff-Quik)
327
B.CSF with C. neoformans.
Rapid Giemsa-like (Diff-Quik)
328
•Detection of encapsulated microorganisms of C. neoformans in CSF.
Colloidal carbon wet mounts (India ink, nigrosin)
329
•The organism is refractory to the particles of ink, and capsules appear as clear halos around the organism against a black background.
C. neoformans in Colloidal carbon wet mounts
330
•Artifacts viz. erythrocytes, leukocytes, and talc particles from gloves or bubbles following a myelogram may
displace the colloidal suspension and mimic yeast (false positive)
331
Determine the test
•When positive in CSF, diagnostic of meningitis. Negative in many cases of meningitis; not reliable. But the sensitivity is (?) followed by lateral flow assay (?) and CALAS (?)
Colloidal carbon wet mounts
97%
94%
74%
332
India ink, basic procedure (1 min.)
1. Addition of 1 drop (?) on a slide
2. Addition of 1 drop
3. Add (?)
4. Observe under the
CSF
India ink
cover slip
microscope 400x
333
Determine the test
•Basic mounting medium for fungi consisting of
Lactophenol cotton blue (LPCB)
phenol, lactic acid, glycerol, and aniline (cotton) blue dye.
334
•Commonly used for the microscopic examination of fungal cultures by tease or tape preparation.
Lactophenol cotton blue (LPCB)
335
makes an excellent permanent stain or fixative for mounting slide culture preparations2.
•LPCB with 10% polyvinyl alcohol (LPCP-PVA)
336
acts as a clearing agent and aids in preserving the fungal structures.
Lactic acid
337
acts as a killing agent
Phenol
338
prevents drying
Glycerol
339
provides color to the structures.
Cotton blue
340
are analogous to cotton blue
Poirrier’s blue and aniline blue
341
LPCB, basic procedure
1. A drop of (?) is added to a glass slide, and a (?) is prepared.
2. Add a (?) and examine at (?)
LPCB solution; tease or tape mount
coverslip; x 100 to x400 magnification
342
Note the crescent spindle-shaped conidia. x 400
Fusarium spp. in LPCB
343
Cells appear as dark-walled and some budding cells x 400.
C. neoformans in LPCP isolated form CSF
344
•Primarily used for the examination of bone marrow, buffy coat, and peripheral blood smears.
Giemsa stain
345
•For the detection of intracellular yeast forms of H. capsulatum and fission yeast cells of Talaromyces marneffei.
Giemsa stain
346
•The fungus is seen as small oval yeast cells, often contained within macrophages, and stains blue; a hyaline halo represents poorly staining cell wall.
H. capsulatum in Giemsa stain
347
•The stain can also be used to visualize trophozoite of P. jirovecii.
Giemsa stain
348
Giemsa stain, basic procedure (15 min.)
1. PBS is prepared on a glass slide and placed in (?).
2. Slide is flooded with freshly prepared Giemsa stain (?)
3. After 5 min. Rinse slide with (?) and air dried. Examine at x 100 to x 400 magnification
100% methanol for 1 min
stock Giemsa stain diluted 1:10 with phosphate buffered H2O
d’H2O;
349
Yeast appears as small, oval cells that stain light to dark blue with a hyaline halo due to the unstained cell wall.
Macrophages from a lymph node filled with H. capsulatum.
350
BAL fluid showing characteristic fission yeast cells of
T. marneffei
351
Detection of bacteria and fungi.
Gram stain
352
Gram positive
Gram negative
yeasts and pseudohyphae
hyphae (septate and coenocytic)
353
may decolorize and appear either Gram negative or stippled
Cryptococcus
354
stain weakly in some instances and exhibit only stippling
Often have orange amorphous material around yeasts
Cryptococcus spp
355
Not all fungi are detected.
Gram stain
356
Filaments of Nocardia stain at least partially acid-fast (pink); Actinomyces and other actinomycetes are negative.
Modified acid-fast
357
Detection of P. jirovecii in respiratory specimens viz. lung biopsy specimen imprints and BAL specimens
Toluidine blue O
358
Stains cyst walls of P. jirovecii reddish blue or dark purple against a light blue background.
Toluidine blue O
359
Cysts are often clumped and may be punched in, appearing crescent shaped.
Toluidine blue O
360
Trophozoites are not discernible.
Toluidine blue O
361
1.Place the air-dried slide in the sulfation reagent (?) for 10 min.
2.Rinse in cold H2O for 5 min., drain, and place in toluidine blue O (?) for 3 min.
3.Rinse in (?), followed by absolute ethanol and then xylene.
4.Examine at (?) magnification.
45 ml of glacial acetic acid mixed with 15 ml of conc’d H2SO4
0.3 g of dye in 60 ml of H2O
95% ethanol
x 100 to x 1000
362
Cytologic stain used primarily to detect malignant cells. But it works well in sputum smears and good for differentiation of dimorphic fungi.
Papanicolaou
363
Depending on the cell type detected, background stains in subtle range of green blue, orange to pink hues.
Papanicolaou
364
Some species of Candida stains gold, while other fungi may not stain at all.
Papanicolaou
365
Mucopolysaccharide stain, not commonly used (more commonly used is mucicarmine stain); like India ink, does not detect all cases.
Alcian blue
366
Useful in visualizing polysaccharide capsule of Cryptococcus that stains blue against a pink background in histological sections of tissue.
Alcian blue
367
The polysaccharide capsule of Cryptococcus neoformans stains turquoise blue; the finer details of the capsule may be better preserved in some specimens when compared with mucicarmine
Alcian blue
368
General-purpose histologic stain. Stains some fungal elements violet to bluish purple in mild to moderate contrast to the lighter background tissue.
Hematoxylin and eosin (H&E)
369
Has advantage of allowing observation of natural pigments of fungi.
Hematoxylin and eosin (H&E)
370
The best stain to demonstrate host tissue reaction and pigment of dematiaceous fungi.
Hematoxylin and eosin (H&E)
371
Fungal cytoplasm is pink and the nuclei are blue.
Hematoxylin and eosin (H&E)
372
Aspergillus spp. and mucoraceous moulds stain well.
Hematoxylin and eosin (H&E)
373
Splendore-Hoeppli phenomenon may be seen with some fungi, including Basidiobolus and Sporothrix.
Hematoxylin and eosin (H&E)
374
Fungi stain pink to red purple, nuclei may be blue depending on the counterstain used.
Periodic acid Schiff (PAS)
375
PAS stains glycogen, so other tissue structures can have a similar appearance to yeast cells.
Periodic acid Schiff (PAS)
376
Demonstrates the double-contoured refractile walls of Blastomyces dermatitidis that may not be visible with GMS stain.
Periodic acid Schiff (PAS)
377
Fungal stain of choice of many dermatopathologists.
Periodic acid Schiff (PAS)
378
Fungal elements including Pneumocystis jirovecii stain gray to black. Background is green.
Gomori methenamine silver (GMS)
379
It also stains Nocardia and other actinomycetes.
Gomori methenamine silver (GMS)
380
Often stains fungi too densely to observe structural details.
Gomori methenamine silver (GMS)
381
Yeast cells and cysts of Pneumocystis may appear similar in size and shape.
Gomori methenamine silver (GMS)
382
Stains mucin
Mucicarmine
383
Stains the capsule of Cryptococcus that appears pinkish red and may also stain the cell walls of Blastomyces dermatitidis and Rhinosporidium seeberi.
Mucicarmine
384
Useful for differentiating C. neoformans (gattii) from other fungi of similar size and shape when found in samples of tissue
Mucicarmine
385
Detection of melanin of dematiaceous fungi and C. neoformans.
Fontana-Mason (FM)
386
Cell walls appear as brown to black with pale pink background.
Fontana-Mason (FM)
387
It was originally thought to stain only dematiaceous fungi and C. neoformans (organisms known to contain melanin or melanin precursors), but it has now been shown to stain variably, but less intensely, A. fumigatus, A. flavus, Trichosporon spp., Fusarium chlamydosporum, and some Mucorales.
Fontana-Mason (FM)
388
Gram stain for bacteria; demonstrates the bacterial filaments of the actinomycetes, eg., Nocardia, Actinomadura.
Brown and Brenn (B&B)
389
Specific method of detecting fungi in body fluids
Fluorescent antibody stain
390
Direct technique: fluorescein-labeled Ab reacts with fungal antigen in cell wall.
Fluorescent antibody stain
391
Indirect technique: unlabeled Ab complexes with fungal antigens. Fluorescein-labeled conjugate reacts with globulins attached to fungal antigens. Cell walls yellow-green.
Fluorescent antibody stain
392
Some use the immunoperoxidase technique. Specific and highly sensitive. Can be used to detect and measure antibodies.
Fluorescent antibody stain
393
Used for the cultivation of ascosporogenous yeasts such as Saccharomyces cerevisiae.
Acetate ascospore agar
394
The potassium acetate formulation has been shown to be a better sporulation medium than sodium acetate.
Acetate ascospore agar
395
Ascospores produced in this medium are visible microscopically after staining Kinyoun carbol-fuchsin acid-fast stain.
Acetate ascospore agar
396
Selective and differential medium used for the isolation of Cryptococcus spp. esp. C. neoformans and C. gattii which is unique in that they produce the enzyme phenol oxidase.
Birdseed agar
397
The breakdown of substrate (Guizotia abyssinica seeds or niger seeds) produces melanin, which is absorbed into the yeast wall and imparts a tan to brown pigmentation of the colonies.
Birdseed agar
398
Colonies of other yeasts are beige or cream in color.
Birdseed agar
399
Chloramphenicol is selective agent that inhibits bacteria and some fungi.
Birdseed agar
400
Creatinine enhances melanization of some strains of C. neoformans.
Birdseed agar
401
Selective and differential medium used for the isolation and differentiation of Candida spp.
Bismuth sulfite-glucose-glycine yeast (BiGGY) agar
402
Nutritive bases include peptone, glucose, and yeast extract.
Bismuth sulfite-glucose-glycine yeast (BiGGY) agar
403
Candida spp. reduce the bismuth sulfite (also acts as inhibitor of bacterial growth) to bismuth sulfide, which results in pigmentation of the yeast colony, with some species, the surrounding medium.
Bismuth sulfite-glucose-glycine yeast (BiGGY) agar
404
C. albicans appear as brown to black colonies with no pigment diffusion and no sheen.
Bismuth sulfite-glucose-glycine yeast (BiGGY) agar
405
C. tropicalis appear as dark brown colonies with black centers, black pigment diffusion, and sheen.
Bismuth sulfite-glucose-glycine yeast (BiGGY) agar
406
Cycloheximide inhibits overgrowth of saprophytic fungi.
Brain heart infusion agar
407
BHI with 10% sheep red cell (enrichment) is used for the cultivation and isolation of all fungi including fastidious dimorphic fungi.
Brain heart infusion agar
408
Chloramphenicol and gentamicin are inhibitors of bacterial growth.
Brain heart infusion agar
409
Adaptation of birdseed agar.
Caffeic agar
410
Caffeic is the biologically active chemical in niger seed that causes the yeast colony of C. neoformans to turn brown.
Caffeic agar
411
Differential medium to distinguish C. neoformans from C. gattii.
Canavanine-glycine-bromthymol blue agar
412
A colony of Cryptococcus is streaked onto the surface of the agar and incubated at 30C for 1 to 5 days.
Canavanine-glycine-bromthymol blue agar
413
C. gattii (serotypes B and C) turns the medium cobalt blue.
Canavanine-glycine-bromthymol blue agar
414
C. neoformans var. grubii (serotype A) and C. neoformans var. neoformans (serotype D) leave the medium greenish yellow.
Canavanine-glycine-bromthymol blue agar
415
Chromogenic, differential and selective medium for the isolation of clinically important yeasts.
CHROMagar
416
Nutritive base is peptone and glucose. Chloramphenicol inhibits bacterial growth.
CHROMagar
417
The medium is available with or without fluconazole, providing additional selection of fluconazoleresistant viz., C. krusei.
CHROMagar
418
More sensitive than SDA and helpful in identifying mixed cultures of yeasts, and it may enhance the rapid assimilation of trehalose by C. glabrata.
CHROMagar
419
Colonies on the medium should be evaluated at 48 h.
CHROMagar
420
Presumptive identification of Cryptococcus, Trichosporon, Rhodotorula spp.
Christensen’s urea agar
421
Urea hydrolysis also facilitates separation of certain dermatophytes viz., T. mentagrophytes and T. rubrum.
Christensen’s urea agar
422
Medium contains urea and phenol red serving as indicator.
Christensen’s urea agar
423
Used for the cultivation and differentiation of T. mentagrophytes from T. rubrum on the basis of pigment production.
Cornmeal agar with 1% dextrose
424
Cornmeal agar with Tween (polysorbate) 80 is used for the differentiation of Candida species on the basis of morphological characteristics.
Cornmeal agar with Tween 80
425
Tween 80 (surfactant) is specifically incorporated in lieu of dextrose for the demonstration of pseudohyphal, chlamydospores, and arthrospores formation.
Cornmeal agar with Tween 80
426
Chlamydospore production is best obtained by subsurface inoculation, or by placing a cover slip over the yeast inoculum, creating a microaerophilic environment— Dalmau method
Cornmeal agar with Tween 80
427
Used for the differentiation of Aspergillus spp.
Czapek-Dox agar
428
Sucrose is the sole carbon source, with sodium nitrate serving as the sole nitrogen source.
Czapek-Dox agar
429
Any bacteria or fungi that can use sodium nitrate as a nitrogen source can grow on this medium.
Czapek-Dox agar
430
Screening medium for the recovery, selection, and differentiation of dermatophytes from keratinous specimens.
Dermatophyte test medium (DTM)
431
Contains cycloheximide that inhibits saprophytic moulds, chloramphenicol inhibits Gram-negative bacteria.
Dermatophyte test medium (DTM)
432
Morphology and microscopic characteristics are easily identified, but pigmentation cannot be discerned because of the presence of phenol red indicator.
Dermatophyte test medium (DTM)
433
Medium is yellow and turns red with growth of dermatophyte.
Dermatophyte test medium (DTM)
434
Used for the isolation and growth of lipodependent Malassezia spp.
Leeming and Notman medium
435
Components of the medium include ox bile, glycerol monostearate, glycerol, Tween 80, and cow’s milk (whole fat).
Leeming and Notman medium
436
Medium may serve as an alternative to SDA because not all species can grow in this medium, eg., M. globosa, M. restricta, M. obtusa which require more complex media for their isolation.
Leeming and Notman medium
437
Selective general-purpose medium used for the isolation of fungi from contaminated specimen.
Littman oxgall agar
438
Crystal violet and streptomycin are the selective agents inhibiting bacteria.
Littman oxgall agar
439
Oxgall restricts the spreading of fungal colonies.
Littman oxgall agar
440
Mycobiotic Remel and Mycosel BD Diagnostic Systems are trade names for a selective medium specifically formulated for the isolation of dermatophytes but also for the isolation of other pathogenic fungi from specimens contaminated with saprophytic fungi and bacteria.
Mycobiotic or Mycosel agar
441
Contains cycloheximide and chloramphenicol.
Mycobiotic or Mycosel agar
442
Inhibited fungi include Candida and Aspergillus spp., mucoraceous fungi, and C. neoformans.
Mycobiotic or Mycosel agar
443
Used to stimulate conidium production by fungi and stimulates pigment production in some dermatophytes.
Potato Dextrose
444
Commonly used with the slide culture technique to view morphological characteristics.
Potato Dextrose
445
The incorporation of tartaric acid in the medium lowers the pH, thereby inhibiting bacterial growth.
Potato Dextrose
446
Consists of pancreatic digest of casein, peptic digest of animal tissue, and dextrose at 4% conc., buffered to a pH of 5.6.
Sabouraud dextrose agar (SDA)
447
Sabouraud dextrose agar (SDA)
448
Emmons modified the original formulation by reducing the dextrose conc’n. To 2% and adjusting the pH nearly to neutrality 6.9 to 7.0.
Sabouraud dextrose agar (SDA)
449
Selective medium that contains:
1.Chloramphenicol
2.Cycloheximide
3.Gentamicin 4.Ciprofloxacin
5.Penicillin and(or) streptomycin
Sabouraud dextrose agar (SDA)
450
Surface area: Large (7,500mm2)
Petri Dish
451
Surface area: Small (1,500mm2)
Test tube
452
Oxygen supply: Good
Petri Dish
453
Oxygen supply: Poor
Test tube
454
Rate of drying: Relatively fast
Petri Dish
455
Rate of drying: Relatively slow
Test tube
456
Security of closure: Poor (lid is easily displaced)
Petri Dish
457
Security of closure: Good
Test tube
458
Probability of dissemination: Relatively large
Petri Dish
459
Probability of dissemination: Relatively small
Test tube
460
Detection of mixed culture: Relatively easy
Petri Dish
461
Detection of mixed culture: Relatively difficult
Test tube
462
Mold forms, most common temperature for incubation
25C to 30C
463
Opportunistic and dimorphic organisms
30C
464
Conversion to yeast phase for dimorphic fungi
25C to 37C
465
< 5 days; Saprobes, opportunistic fungi, yeast
Rapid growers
466
6 - 10 days; Subcutaneous & opportunistic fungi, dermatophytes
Intermediate growers
467
> 11 days; Systemic & subcutaneous fungi
Slow growers
468
Appears leather-like, or waxy little mycelium, seems to merge with the agar
Glabrous
469
Resembles plush or suede, short aerial hyphae of equal length
Velvety
470
Resembles colonies of other Staphylococcus spp. (formerly CoNS), “bacteria like” but more dry and dull (waxy-pasty). No aerial mycelium, with a delicate fringe around the colonies in BAP
Yeast like
471
Wooly or “Floccose”, large quantities of long aerial hyphae that becomes entangled and may fill the entire petri dish
Cottony
472
Powdery due to heavy conidiation or sporulation, has even hyphae and abundant conidia
Granular
473
Presence of radial groves from the center of the culture toward the rim
Rugose
474
Random folds (long, short, parallel at right angles or combination)
Folded
475
Have central depression (concavity) surrounded by raised edges. Resembles S. pneumoniae colonies
Crateriform
476
Have many warts or rough knobs on the surface
Verrucose
477
Brain-like convolutions
Cerebriform
478
1.Place 1-2 drops of the mounting fluid (eg., LCB) on the slide. 2.Gently remove a portion of the colony from the agar and place it on the slide.
3.Using two teasing needles, tease the strands apart.
4.Place a coverslip and examine under a microscope
Tease mount
479
1.Wrap a piece of clear cellophane tape around a tongue blade, with the sticky side out.
2.Touch the colony with the sticky tape.
3.Lay the tape, sticky tape down on a drop of mounting fluid on a slide.
Scotch tape method
480
1.Cut a square block (1 cm2 or 15 mm2) form an agar (eg., SDA, Cornmeal, PDA), then transfer on a sterile glass slide.
2.Inoculate all sides of the agar block, then cover with a cover slip. (22 x 22 mm).
3.Place the set-up in a moist (humidified) chamber (wet house) and incubate until growth appears.
4.Prepare a sterile glass slide with 1-2 drops of mounting fluid (LPCB).
5.Carefully remove the coverslip from the block by lightly twisting. 6.Lower the coverslip carefully onto the mounting fluid.
7.Set aside and examine under a microscope.
Slide culture (syn. Microculture)
481
1.Make one streak of a young, actively growing yeast down the center of the area (do not cut the agar); make three or four streaks across the first to dilute the inoculum.
2.Cover with a 22-by-22-mm coverslip.
3.Incubate at room temperature, in the dark, for 3 days.
4.Examine by placing the plate, without its lid, on the microscope stage and using the low-power (X100) and high-dry (430) objectives. The most characteristic morphology (especially the terminal chlamydospores of Candida albicans) is often found near the edge of the coverslip.
Dalmau Method
482
Conidia and spores form a fresh culture are washed off in sterile water and placed in labelled vials
Storage in wate
483
Labelling glass tubes with a screw-capped and placed in a freezer (-70C)
Freezing
484
Layering an entire slant with mineral oil, capping the tube tightly and storing at room temperature
Mineral oil overlays
485
Freeze drying with the use of special equipment (lyophilizer)
Lyophilization
486
Several immunological techniques have been used to detect antibodies during fungal infections
1.Immunodiffusion (ID)
2.Countercurrent immunoelectrophoresis (CIE)
3.Enzyme-linked immunosorbent assay (ELISA) 4.Complement-fixation test (CFTs)
5.Fluorescent-enzyme immunoassay (FEIA)
487
CIE, FEIA, EIA
Aspergillus IgGAspergillus IgG
Ab detection is helpful in diagnosing allergic bronchopulmonary aspergillosis (ABPA), aspergilloma, and Chronic cavitary aspergillosis
Aspergillus spp.
488
Commercial kits
Antigens (eg., A Ag, WI1 Ag)
Limitation: lack of sensitivity due to crossreactivity with dimorphic fungi
Blastomyces spp.
489
EIA
Mannan Ag Candida IgG, IgM, & IgA
Sensitivity 62% Specificity 53%
Candida spp.
490
EIA
Cryptococcus IgG, IgM, IgA Ag: Glucuronoxylmannan
Ab detection can be a marker for reactivation of disease in solid organ transplant recipient
Cryptococcus spp.
491
A polysaccharide present in the cell wall of many fungi viz., Aspergillus, Candida, Fusarium with little or none on the following viz., Cryptococcus, Blastomyces, Mucoraceous molds.
1,3--D-glucan detection
492
Found in the blood of patients with invasive fungal infections, and its ability to activate factor G of the horse-shoe crab coagulation pathways that allows it to be measured and quantified.
1,3--D-glucan detection
493
Has the ability to detect organisms that are present in small numbers of that cannot be cultured.
Nucleic acid testing
494
Removal of the risks assoc. with culturing category 3 (BSL 3 fungi)— especially for H. capsulatum
Nucleic acid testing
495
Decrease time for identification of the agent, & the potential to detect anti fungal resistance in primary clinical specimens
Nucleic acid testing
496
Offers the potential to identify pathogens in biopsy samples that have been formalin-fixed and even wax-embedded.
Nucleic acid testing
