Introduction to Clinical Mycology: Part 1 [Hot Topic]

Welcome to Mayo medical laboratories
hot topics.

These presentations provide short
discussion of current topics

and may be helpful to you
in your practice.

Our speaker for this program is
Dr. Glenn Roberts

a professor of laboratory medicine
and pathology and microbiology

as well as a consultant in the division
of clinical microbiology at Mayo Clinic

in Rochester Minnesota.

Dr. Roberts provides a
four part introduction to

clinical mycology including culture
and identification

of organisms encountered
in the clinical practice.

This is part one in the series,
thank you Dr. Roberts.

Thank You Sharon for that introduction.

I have nothing to disclose.

Introduction to clinical mycology.

This is the first in a series
of four presentations on

clinical mycology.

Today is part one.

Discusses what the clinical laboratory
does to make the diagnosis

of a fungal infection.

Discusses the classification of fungal
infections in the fungi related to them.

It also discusses the general features
of the fungi their importance in the basic

morphological features of the molds.

I think we periodically need to think
of why we're working in the field

that we are in
and that is to support patient care.

And a lot of activities go on
behind the scenes that

we're not aware of for the most part.

And one is that a patient with
some symptoms of something

whether it's a fungal
infection or something else,

and for our purposes
we'll say it's a fungal infection

"come in with signs and
symptoms of infection".

The clinician has to decide what
he or she suspects might be the problem.

Ask pertinent questions,
like a history of travel.

And then investigate the immune status
of the person,

because fungal infections take advantage
of people who are immunosuppressed.

They then have to decide
if they're going to culture something,

and they select appropriate specimen
for that.

They have to order a direct microscopic
examination if they think it's appropriate,

So that we can make
a rapid diagnosis in some instances.

And then sometimes it even
goes a lot further than that,

where they involve in basic procedures
like bronchoscopy or

fine needle biopsy or something like that.

And other things that can be done
or serologic tests for antigen antibody

and some metabolites
of some of these fungi

that assist in making a diagnosis prior
to maybe doing an invasive procedure.

If a culture is ordered
and something grows,

they will perhaps order
as and a fungal susceptibility test.

And then the laboratory sometimes is
involved interpretation of those results,

whether it's a serologic test or whether
it's anti fungal susceptibility test.

And then the most important thing
and it will reemphasize

this again in just a moment
is communication with the physician.

The laboratory must communicate with
a physician and not be afraid to call

the condition up and give them results
of patient care

for about the patient result,

so that we don't delay therapy
for these patients.

And oftentimes we have to also go back
and communicate with others

in a laboratory to make sure
we have all this information straight.

Well what is the laboratory involved
with?

Well oftentimes we're to ask
questions about a collection

of specimens by the clinician and we need
to be aware of what those rules are.

We need to be proficient at the direct
microscopic examination of specimens.

We know about culturing,
we identify the yeast and molds

in the laboratory.

We either refer out or do
antifungal susceptibility tests

and fungal share logic testing.

We're involved in reporting results
and again here we are,

we're communicating with physicians again.

And this is the key part of what
we do is communication with our

colleagues and with our physicians.

Well what are fungal infections;
what are some general features?

Well first of all these
fungi may affect normal

and immunocompromised patients.

Many of them are chronic and duration
they last for years like a ringworm

some things like that,

but in the immunocompromised
patient these can be acute infections

and progressed very quickly to death.

They're not transmitted from
patient to patient

they are not susceptible
to the usual antibacterial agents

that we know about.

However they are susceptible
to several groups

of antifungal drugs called
polyenes, azoles, and some others.

And the therapy for treating
fungal infections

is not easy for the patient -
very difficult.

Side effects are substantial
and we need to make sure that we make

a accurate diagnosis before these patients
are placed on therapy.

And again these infections
may range from being

just chronic type things to some that
are acute and progressed to rapid death.

And so we need to be
thinking about a rapid

reporting of results as we do our work.

Well we try to classify
fungal infections into

groupings and it's hard to do,
because it's an artificial thing.

But we divide them up into
superficial infections, subcutaneous,

systemic,
and opportunistic infections.

The superficial infections
you can remember by

just thinking about the fact that
they involve the keratinized tissue.

The hair, the skin, and the nails,
and that your matter fights

are good examples
of those and some others.

Subcutaneous infections
involve the skin and the contiguous

subcutaneous tissues
that includes the lymphatic vessels.

And most of the infections that
are involved in subcutaneous tissue

are acquired by trauma to a site
using the extremity.

Systemic infections
can involve any organ system

and generally they're caused by
certain groups of fungi that

have specific geographic niche
at an environment.

And they live in certain
locales within the world,

and we know where those are
and so we ask appropriate questions

when the clinician does anyway
ask some appropriate questions.

But these are organisms that
are involved in this

actually can infect any organ system,

and actually do that
and the patients may be

totally asymptomatic and unaware of this.

Or they may be very very ill
if they happen to be a know compromised.

And then the last
group is very important one,

opportunistic fungal infections.

These are infections caused by fungi that
normally don't cause disease in humans.

Their environmental flora.

And if we have a breakdown
of our immune system,

then we - and we happen to be
in the wrong place at the wrong time.

Then we acquire these infections
and they progress

very quickly particularly
in transplant patients,

and so we need to be aware
of what organisms cause these infections.

The hardest part of mycology
is the terminology, It's a language.

And I'm just going to go through
very quickly here some of the names,

so you can see them.

Superficial infections
of dermatophytes involved primarily

causing these infections.

Another northern
organism in however

that does cause infection
and causes the tinea versicolor

is called Malassezia Furfur.

Dermatophytes belong
the three groups

Trichophyton, Microsporum,
and Epiduralphyton.

And these are just here to show
you what the names look like.

Subcutaneous infections: Sporotrichosis,
Mycetoma, Chromoblastomycosis,

and Phaeohyphomycosis.

It caused by Sporothrix schenckii,
Pseudalleschria boydii,

Phialaphora verrucosa,
Cladophialophora carrionii respectively.

And you can see the names
get a little complicated sometimes

they're not that hard once you learn.

Terms of systemic infections:
Histoplasmosis, Blastomycosis,

Coccidioidomycosis,
and Paracoccidioidomycosis.

or the infections
that are grouped into

this particular heading,

and they're extra to another one
which is not listed here

and I mentioned it to you in a minute.

The fungi that are involved in causing
those infections Histoplasma capsulatum,

Blastomyces dermatitidis,
Coccidioides immitis,

Paracoccidioides brasiliensis.

An one called Penicillium marneffei
that causes Penicilliosis

in places like Cambodia,
Vietnam, Thailand.

We don't see it in this country
very often, but we do see it.

Opportunistic infections examples
of Candidiasis, Cryptococcosis,

Aspergillosis, Zygomycosis,
and a whole lot more.

The list could go on and on
because we see organisms

that we have never thought
would cause disease.

Cause disease in patients here
and cause substantial infection

and often as leads to death.

Some of the organisms related to that
Candida albicans, Cryptococcus neoformans,

Aspergillus, rise up as Mucor,
and a number of other

of the Mucorales Fusarium, Acremonium,
Trichosporon and many others.

Let's talk about the fungi and -
and in general here just a moment that

they are eukaryotic cells.

They have a cell wall.

They have a cell membrane
and the cell walls made up

of poly-n-acetylgucosamine,

and the cell membrane contains
ergosterol in contrast to maeín cells

which contains cholesterol
and the reason for that

I mention in this is that
there are certain of these

anti-fungal agents that actually bind
to the cell membrane

that has ergosterol in it.

The heterotrophic;
They break down an organic matter

to get energy sources for
themselves decaying matter.

They are not susceptible again
to the usual antimicrobials.

They reproduce sexually
or asexually most

of the time in the laboratory,

we see the result of asexual
reproduction with spores.

They may be monomorphic
which means they have one

form a yeast or a mold form,

they may be dimorphic
and that is if they have a yeast

and a mold form or another type form.

And then some of them actually
produced more than two or three forms,

and they're called polymorphic.

So reproduction by these
is used by asexual spores

and that's what we see all the time
when we looked at these organisms

underneath the microscope.

Examples of the fungi that
you'll be familiar with our yeast

and molds which we'll talk about.

Mushrooms, puffballs,
shelf fungi, the morels,

which I'm sure some of you have collected.

Plant pathogens and animal pathogens.

And some of these are
very substantial things

that are involved in causing disease
of plants and animals.

One of the things that the fungi are very-

have a very important role in doing
is keeping the carbon cycle going,

they break down organic
matter decompose it into carbon.

They cause disease
and trees for example a Dutch elm disease,

which is a devastating thing
for a population of trees

and that we lose them all
whenever they get Dutch elm.

Crop diseases;
Wheat, corn, barley.

Corn smut is one of those examples of that
and we actually have seen patients

who have had disease
caused by corn smut.

Wood and timber degradation;
If you look at wood you'll see that

there's some purple to blue
material sometimes on there

where the wood gets broken down by
these fungi and it damages it.

Another very important thing is
these fungi can cause spoilage

of food particularly grain.

And developing countries
where grain as a primary staple.

The grain will get infected with a fungus
and produce a toxin called aflatoxin,

if it happens to be
infected with aspergillus.

and actually caused substantial disease
in these patients

and oftentimes liver cancer.

And where would we be without fungi
and the brewing and baking industry?

We would be nowhere,
because the yeast

of the things
that cause fermentation.

So their play a major role in that.

Well what are these fungi in general?

We mentioned their heterotrophic.

They break down organic matter,
they have chitin in the walls,

ergosterol in the cell membrane,
they have an organized nucleus,

they reproduce by asexual spores
again most often.

And if it happened to be a mold
they produce hyphae,

which you can see on
this left-hand photograph.

They're nothing more than filaments
that look like garden hoses

underneath the microscope.

And if it happens to be a yeast
they produce single cells that

reproduce by budding,

and oftentimes a
few more structures in there.

So just to give you an idea
of some of the terms.

Some more language
if you will to learn.

But we have terms that we use to help us
describe what we see under the microscope

to make it a little easier
for others to understand.

Well if you look at a mold
on a culture plate

you'll see the colony there,

a fluffy colony and our laboratory
we call them fuzzies.

If you look underneath
the microscope

you'll find out
they're made up of these hyphae,

these are filaments
that have parallel walls

you look like small garden hoses.

The collective name
for the colony is mycelium,

and I don't think many people use
the term mycelium anymore

but it's there for you to see.

The hyphae; these garden hose
type structures may be divided up

in compartments
by some structures called septa,

and so we talked about septate hyphae.

And we talked about non-septate hyphae
which don't have any of these

division these compartments in them.

And sometimes we just refer
to non septate hyphae as pauciseptate,

because certainly these fungi that are -
that are thought to be non septate

have a few of these separations and I'll
show you shortly here.

Spores; most of the fungi that we deal
with produced spores

that are called conidial.

And they're produced on
specialized structures

whether it's a short to long stalk
or really elaborate structure

called a conidiophore.

Some of the spores are small,
some of them are large.

And so we define them by being
macroconidia or microconidia.

And sometimes these spores
or hyphae may be pigmented

or they may not be
and that leads us to different groups.

Well just some more terms here
the hyphae we talked about

these are structures that
make up with a mold colony.

The septa are the cross walls
that break down the hyphae

into compartments.

Non-septate means that
they're lacking septa

like we said while ago.

And hyaline is another term
you haven't talked about yet

that is and that the hyphae may be
non-pigmented.

In other words if you look at them
underneath the microscope

without any stains at all
they'll be clear.

If you stain them with a dye
like we do in the laboratory

lactophenol and on blue,

you can see they'll turn blue
with it with a dye.

But they still don't have
any definable pigment to them,

so they're called hyaline.

And then we have other
fungi that are dematiaceous,

and these fungi are the ones
that contain a dark pigment.

Either kind of a chestnut brown pigment
or very black.

They belong to a whole different group
of fungi sometimes some

of which are very difficult to identify.

And then there are a lot of structures
that we have to deal with,

and some of them are just nonspecific kind
of things that don't tell you

anything about the organism.

And one of those is called a
Chlamydoconidium.

It's a big round spore
found right up within

the hyphae strand
or on the end of the hyphae strand.

And it's the primary function is
to protect the organism so that if it gets-

becomes under adverse conditions
it'll round up and form these spores

so they can survive.

We say are non specifically in
the lots of organisms.

And then the Conidia.

The conidia are those asexual spores
you mentioned produced by

molds that have septae.

And the reason I mentioned with septae
is that there are a whole group

of fungi that do not produce asexual spores
and have type septate hyphae.

They happen to have non septate hyphae,
and the spores are produced

in a different way
and we'll talk about that.

This is a typical mold colony.

Colonies are things that we see
that we have to work from

to make them out in the laboratory
to be able to identify the organism.

However looking at a culture plate
sometimes gives you a little information,

but many times doesn't help you at all.

The colonies can
be multicolored.

They can be white, off-white, brown, tan,
green, yellow, pink, brown, black,

there could be all sorts of colors.

You may get a ballpark idea
of what the organism is,

but it's not going
to tell you specifically what it is.

This is another one or you can see the
colonies are smooth adherent to the augur.

And they have different morphological forms
when you start looking at these colonies.

They look either very fluffy
or very adherent to the augur,

and other appearances as well.

So you look at molds you'll be surprised
at the variety of things that you see,

and you have to kind
of you have to with experience

you can tell sometimes
which are the ones

you think are going to be important.

Now this is the example of the hyphae.

This is an electron micrograph
where you can see

these garden hose
structures I told you about.

And there is a septum
that cross wall that divides

that organism up.

And there'd be another
septum probably down below it there,

and it allows that organisms
to be compartmentalized.

So that if it happens to be a break apart.

Each compartment can then grow,
and start a new colony.

So it's a matter - it's kind

of a motor survival
for these organisms.

This is what it looks like
under the microscope

when you really look at it.

You can see the septum there
to the right of the arrow

that divides it up.

And they're not usually
so hard to see as this one,

but sometimes they are.

This is an organism that is Dematiaceous,
in other words it's pigmented.

And one of the things that
you do in the laboratory is

to recognize these molds sometimes by the-

When you look at the way
the spores are produced

you look at the spores to recognize
the size and shape

of the spores and so on.

And you can look at this one and say

"ah I know what this is
this is by Polaris.",

because of the features that
you'll be familiar with as we go through

the few other sessions.

But the arrow here shows those septae,

and you can see the this hyphal strand
at the bottom left hand corner

is divided up into compartments.

So that each one of those
is an separate unit

and can survive and grow
and produce a new colony.

And then the image here
is one of non septate hyphae.

These are very large. They're not
compartmentalized at all.

And when the laboratory process
is something that's suspected

to have one of these organisms
that has non septate hyphae.

If you grind it up like people do
in the laboratory,

that hyphal strand will pop open.

All the cytoplasm will
leak out and it'll die.

So what you have to do is be very careful.

You end up having
to not grind the culture up,

you end up having to cut
pieces of whatever it is.

Tissue or whatever it is,
so that you can allow

survival for this organism.

But these hyphae are non septate,
and occasionally

they will have septations and we call
them pauciseptate.

This completes part 1.

The introduction of Clinical Mycology.

future presentations will be part 2:

Reviews the basic structures
of molds and yeast.

Presents a brief introduction
to the direct microscopic examination

of clinical specimens
and also media

that are useful for culture.

Part 3 presents specific
information on the culturing

and incubation of cultures
for the off more recovery of fungi.

And part 4 presents the methods
for identification of fungi,

primarily molds,
and some helpful hints

for working within
the clinical laboratory.