Introduction to Clinical Mycology: Part 1 [Hot Topic]

welcome to mail 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
moles 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 as 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 ser logic test or whether
it's anti fungal susceptibility test and
then the most important thing and it
will resize 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 library
the yeast in moles 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
- the usual antibacterial agents that we
know about
however they are susceptible to several
groups of antifungal drugs called poly
means a zoals 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 an
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 or 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 in the College tinea
versicolor is called malassezia furfur
dermatophytes belong the three groups
trichophyton microsporum and epidural
fighting and these are just here to show
you what the names look like
subcutaneous infections sports are kosis
my Sonoma krummel blastomycosis and fail
- mycosis it caused by sport through
kinky I sued Alisher boy he fell off her
varicose and Claro file off her carry-on
irrespectively 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 para
coccidioidomycosis 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 funds either involved in causing
those infections Histoplasma capsule
item blastomyces dermatitidis
Coccidioides immitis para Coccidioides
brasiliensis in one called penicillin
Martin Fe that causes penisula sis and
the 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 in the elephant as leads to
death some of the organisms related to
that Candida albicans Cryptococcus
neoformans Aspergillus rise up as mu
core and a number of other of the mucous
Fusarium acrimony amicus Poron and many
others let's talk about the fungi 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 an acetic osa mean
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 rostral 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 a sexually most 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 that they have a
yeast and a mole 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 puff balls shelf funds
out of the morels which I'm sure some of
you have collected plant pathogens and
animal pathogens and some of these are
very substantial things they are
involved in causing disease of plants
and animals one of the things that the
funds I 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
there 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
wooden 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
it's fungi and it damages it another
very important thing is these fungi can
cause spoilage of food particularly
grain and then 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 a spore
jealous 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 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 heterotroph
as they break down organic matter they
have chitin in the walls
yeah Burgas rot in the cell membrane
they have an organised nucleus they
reproduce by a suction source again most
often and if it happened to be a mold
they produce high P which you can see on
this left-hand photograph they're
nothing more than filaments that look
like garden hose is 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 languages 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 mole on a culture plate you'll
see the colony they're a fluffy colony
and they're our laboratory we call them
fuzzies if you look underneath the
microscope you'll find out they're made
up of these high P 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 mean it
might see them anymore but it's their
sory to see the hyphy these garden hose
type structures may be divided up in
compartments by some structures called
septic and so we talked about septate
high P and we talked about non septate
hi people which don't have any of these
division these compartments in them and
sometimes we just refer to non septate
high here's policy septic because
certainly these fungi that are that are
thought to be non subset
have a few of these separations and I'll
show you shortly your spores most of the
funds I 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 a really elaborate structure
called a Canadian for some of the spores
are small some of them are large and so
we define them by being macro conditio
or micro Canadia and sometimes these
spores are - may be pigmented or they
may not be and that leads us to
different groups well just some more
terms here the - we talked about these
are structures that Mick with a mole
colony the septa are the cross walls
that break down the - into compartments
non septate means that they're lacking
septa like we said while ago and
Highland is another term you haven't
talked about yet is the hyphy 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 say them with a dye like we
do in the laboratory lacto phenol 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 Highland and then we have other
fungi that are DeMatteis and these funds
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
chlamydia Canadian it's a big round
sport found right up within the - strand
or on the end of the - 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 forests so they can
survive we say are not specifically in
the lots of organisms and then the
Canadian the Canadian of the say sexual
spores you mentioned produced by moles
that have septa and the reason I
mentioned with septa is that there are a
whole group of fungi that do not produce
asexual spores and
type septate - they happen to have non
septate - and the spores are produced in
a different way and we'll talk about
that this is a typical mole 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 moles you'll be surprised
at the variety of things that you see
you have to count the effort with
experience you can tell sometimes which
are the ones you think are going to be
important now this is the example of the
high P this is an electron micrograph
where you can see these garden hose
structures I told you about and there is
a septum the 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
DeMatteis in other words it's pigmented
and one of the things that you do in the
laboratory is to recognize these moles
sometimes by the when you look at the
way the spores are produced you look at
the scores 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
septic and you can see the
this Highfill strand at the bottom left
hand corner is divided up into
compartments so that each one of those
is an exception and can survive and grow
and produce in your colony and then the
image here is one of non septate havi
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 cefotetan - if you grind it
up like people do in the laboratory that
Highfill 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
high PR non septate and occasionally
they will have cept Asians and we call
them policy sceptic this completes part
1 the introduction of clinical mycology
future presentations will be part 2
reviews the basic structures of moles
and yeast presents a brief introduction
to the direct microscopic examination of
clinical specimens and also media that
are useful for culture part 3% 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 moles and some helpful hints
for working within the clinical
laboratory