SIMIT 2010
Gestione pratica delle infezioni fungine
in terapia intensiva
Epidemiologia, fattori di rischio
e diagnosi delle infezioni
fungine in terapia intensiva
Nicola Petrosillo
UOC Infezioni Sistemiche e dell’Immunodepresso
Istituto Nazionale per le Malattie Infettive
“Lazzaro Spallanzani
Invasive Aspergillosis (IA) in the ICU
•The bulk of literature about IA involves
patients with classic risk factors for IA, such
as prolonged neutropenia and hematopoietic
stem cell transplantation.
•However, a broad group of patients who are
admitted to ICUs may also be susceptible to
these infections.
IS IA A PROBLEM IN THE ICU?
•Estimates about the incidence of IA among critically ill
patients are sparse and variable.
•For various reasons, figures about the true incidence of
IA are difficult to generate:
1. discrimination between colonization and
infections remains challening;
2. few istitutions perform autopsies routinely;
3. characteristic radiological signs of IA are usually
absent in the nonneutropenic ICU patient.
4. diagnostic utility of non-culture based
microbiological tools has not been properly
validated in the nonhematology ICU population;
5. EORTC/MSG guidelines were not designed for
patient categories other than patients with cancer
and patients who have undergone bone marrow
transplantation
Retrospective autopsy-controlled
studies
•A study sought unsuspected causes of death in a ICU
revealed that, among 100 autopsies, there were 15 cases
of IA, of which 5 were missed before death.
Roosen J et al. Mayo Clin Proc 2000; 75:562–7.
•127 (6.9%) of 1850 hospitalized patients had
microbiologic or histopathologic evidence of
aspergillosis during their ICU stay, including 89 cases
(70%) in which there was not an underlying hematological
malignancy.
•The observed mortality rate of 80% was much higher
than the mortality rate predicted on the basis of the
Simplified Acute Physiology Score II (48%).
Meersseman W et al. Am J Respir Crit Care Med 2004;170:621–5
Other epidemiological data on IA in the ICU
•IA prevalence in pts with septic shock  0.3%
(Vandewoude KH et al. Crit Care 2006; 10:R31).
• High prevalence of IA (pathologic and/or microbiologic
evidence of aspergillosis) in a cohort of patients with
severe hospital-acquired pneumonia who had been
admitted to the ICU 13 (19%) of 67 episodes of IA in the
ICU
(Valles et al. Intensive Care Med 2003; 29:1981–8).
•During a 6-year period, Cornillet et al. found that a mean
number of 15 patients per year received a diagnosis
of IA; approximately one-half of these patients were in
the ICU.
(Cornillet A et al. Clin Infect Dis 2006; 43:577–84).
WHO IS AT RISK OF
DEVELOPING IA
IN THE ICU?
Meersseman W et al. Clin Infect Dis 2007; 45: 205-16
Which diseases are
associated to IA in
the ICU?
COPD AND ASPERGILLOSIS
• In a review of 50 studies, COPD was the underlying
condition in 26 out of 1,941 (1.3%) patients with
Lin SJ et al. Clin Infect Dis 2001; 32: 358-66
aspergillosis
• In one large study, 9% of 595 patients with IA suffered
from pulmonary disease Patterson TF et al. Medicine 2000; 79: 250-60
• Steroids are believed to play a role in the emergence
of IA, and some authors have investigated the
correlation between the daily dose of corticosteroids
and the probability of developing IA Leav BA et al. N Engl J Med
2000; 343: 586
Samarakoon P et al. Chronic Resp Dis 2008; 5: 19-27
Tools for diagnosis of invasive
aspergillosis
and their applicability in the ICU
CT
Halo sign  sign arrives too early (5 days
before the onset of disease).
Not specific for Aspergillus spp
(also other molds)
Crescent sign obscured by atelectasis,
ARDS, and/or pleural effusion.
CT often is not feasible in a patient with a high
fraction of inspired oxygen
Balloy V et al. Infect Immun 2005; 73: 494-503
Bulpa P et al. Eur Respir J 2007; 30: 782-800
Bulpa P et al. Eur Respir J 2007; 30: 782-800
Bulpa P et al. Eur Respir J 2007; 30: 782-800
Bulpa P et al. Eur Respir J 2007; 30: 782-800
Histopathologic evidence
[acutely branching (45°), septated hyphae
mainly in lung tissue]
It is a global standard for ICU
Roosen J et al. Mayo Clin Proc 2000; 75: 562-7  100 IA
Meersseman W et al. Am J Respir Crit Care 2004; 170:621-5 129 IA
However, biopsies often are not feasible in
patients with thrombocytopenia or a high fraction
of inspired oxygen
Culture
(Sabouraud agar)
•Poor sensitivity and specificity.
•Isolation of the species takes several days.
•50% of cases are missed on the basis of culture
•and microscopy findings.
•Discrimination of colonization versus invasive
•disease is difficult
•Positive predictive value increases with increased
immunosuppression.
Direct microscopy
• PAS, Grocott stain, calcofluor visualization
of hyphal elements (not only Aspergillus spp),
rapid test.
• Same problems of Culture
Galactomannan serum assay
(threshold, 0.5-1.5 ng/mL)
• Tested mainly for haematologic non-ICU patients
• False positive (Pip/Taz)
• In the nonneutropenic critically ill patients, BAL
may perform better than serum
Meersseman W et al. Am J Respir Crit Care Med 2008; 177:27-34
Meersseman W et al. Am J Respir Crit Care Med 2008; 177:27-34
Meersseman W et al. Am J Respir Crit Care Med 2008; 177:27-34
•Paziente con COPD in terapia cronica steroidea.
•Ricovero in ICU per esacerbazione di COPD ed
insufficienza respiratoria.
•BAL colturale positivo per Haemophilus influenzae
e negativo per funghi.
•Galattomannano serico negativo, ma positivo su
BAL.
•Trattato con terapia antiaspergillare, ma deceduto.
•Autopsia aspergillosi limitata al polmone
Meersseman W et al. Clin Infect Dis 2007; 45: 205-16
PCR
• PCR holds promise for patient with haematologic
malignancy, but has not been systematically studied
for the diagnosis of IA in the ICU
• In the nonneutropenic, critically ill patient, BAL may
perform better than blood
• PCR of respiratory secretion specimens as a modality of
surveillance is an interesting topic of research
Beta-(1,3)D-glucan
• hampered by false positive results (albumin,
wound gauze, hemodialysis, bacterial infection).
• Not specific for Aspergillus spp; also present in
yeasts and bacteria
• May be useful as a negative predictor of fungal
infection
What is the meaning of
Aspergillus colonization?
Khaasawneh F et al. J Crit Care 2006; 21: 322-7
Bouza E et al. J Clin Microbiol 2005; 43:2075-9
Candida in ICU
Suspected invasive candidiasis in the
ICU
An invasive Candida infection is typically considered
when a febrile nonneutropenic patient in the ICU who has
some risk factors for Candida infection
•vascular access,
•administration of TPN,
•the receipt of corticosteroids,
•therapy using broad-spectrum antimicrobial agents,
•recent surgery,
•a prolonged stay in the ICU,
•colonization with Candida
fails to respond to broad-spectrum antibacterial therapy
De Pauw BE et al. Clin Infect Dis 2008; 45: 1813-21
Rapid progression of IC: a reason for
suspecting Candida infection in the ICU
Day to start of fluconazole
Garey KW et al. Clin Infect Dis 2006; 43:25-31
Delay in starting antifungal therapy is an independent
determinant of hospital mortality in pts with candidemia
Morrell M et al. Antimicrob Agents Chemother 2005; 49:3640-5
Candida in the ICU
- epidemiology•
The past two decades have marked a dramatic rise
in the frequency of infections caused by Candida
species.
•
1% to 8% of patients residing in the hospital
develop invasive candidiasis versus 10% of
patients residing in ICUs
(Eggimann P et al. Lancet Infect Dis 2003; 3:685–702).
•
Invasive candidiasis accounts for up to 15% or 30%
of all nosocomial infections in critically ill patients.
(Eggimann P et al. Lancet Infect Dis 2003; 3:685–702; Magnason S et
al. Acta Anaesthesiol Scand 2008;52: 1238–45)
Candida in the ICU
- epidemiology-
•About 80% of cases of candidemia arise from or evolve
in the presence of a vascular access, including access
related to central venous catheters, hemodialysis
catheters, peripherally inserted central catheters, and
implanted ports.
•An estimated 33% to 55% of all episodes of candidemia
occur in patients in the ICU, and the associated mortality
rates range from 5% to 71%.
Ben-Ami R et al. J Clin Microbiol 2008;46:2222–6
Bouza E et al. Int J Antimicrob Agents 2008;32(Suppl 2):S87–91.
Candida in the ICU
Risk factors
•Two main factors predispose to infections with Candida
spp.:
- colonization of skin and mucous membranes with
Candida and
- alteration of natural host barriers (wounds, surgery,
and insertion of indwelling intravascular and urinary
catheters).
•The gastrointestinal tract, the skin and the urogenital
tract are the main portals of entry for Candida infections.
•Since 1994, colonization by Candida spp. has been
established as a major risk factor for invasive candidiasis
[Pittet D et al. Ann Surg 1994; 220:751-758].
Candida in the ICU
- Risk factors-
Risk factors for candidemia in patients in the ICU
include:
•the use of intravascular catheters,
•parenteral nutrition,
•prior abdominal surgery,
•the use of broad-spectrum antibacterial therapy,
•the use of corticosteroids,
•acute renal failure,
•a prolonged stay in the ICU,
•and Candida colonization, particularly if it is
multifocal.
Bouza E et al. Int J Antimicrob Agents 2008;32(Suppl 2):S87–91.
Leroy O et al. Crit Care Med 2009; 37: 1612-18
Bassetti M et al. BMC Infect Dis 2006; 6: 21
Bassetti M et al. BMC Infect Dis 2006; 6: 21
Bassetti M et al. J Antimicrob Chemother 2009; 64: 625-29
30
25
#
20
C albicans
C parapsilosis
C glabrata
C tropicalis
other Cand
15
10
5
0
1999 2000 2001 2002 2003
Bassetti M et al. BMC Infect Dis 2006; 6: 21
Epidemiology of Candidaemia in Europe: Results of 28-Month European
Confederation of Medical Mycology (ECMM) Hospital-Based Surveillance
Study
Tortorano AM et al. Eur J Clin Microb Infect Dis 2004; 23: 317-22
• 76 pts with Fl-R C glabrata BSI
• 68 pts with Fl-S C glabrata BSI
• 512 controls
Lee I et al. Arch Intern Med 2009; 169: 379-83
In vitro susceptibility to fluconazole of Candida species in
patients naive for azole agents and in previously exposed to
azole agents
S= susceptible S-DD= susceptible dose dependant
R= resistant
Leroy O et al. Crit Care Med 2009; 37: 1612-8
Epidemiology of Candidaemia in Europe: Results of 28-Month European
Confederation of Medical Mycology (ECMM) Hospital-Based Surveillance
Study
Mortality rates by etiological agent
45
40
35
30
25
20
15
10
5
0
C
alb
C
ica
ns
gla
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Tortorano AM et al. Eur J Clin Microb Infect Dis 2004; 23: 317-22
Prospective, observational, multicenter, French study conducted from October
2005 to May 2006
•Among the 136 patients analyzed, 78 (57.4%) had
candidemia caused by C. albicans.
•These patients had
-earlier onset of infection (11.1 ± 14.2 days after ICU
admission vs. 17.4 ± 17.7, p = 0.02),
-higher severity scores on ICU admission (SOFA: 10.4 ± 4.7
vs. 8.6 ± 4.6, p = 0.03; SAPS II: 57.4 ± 22.8 vs. 48.7 ± 15.5, P
= 0.015), and
-were less often neutropenic (2.6% vs. 12%, p = 0.04)
than patients with candidemia due to non-albicans
Candida species.
Leroy et al. Critical Care 2010, 14:R98
Role of Colonization
Candida Colonization of the Respiratory Tract and Subsequent
Pseudomonas Ventilator-Associated Pneumonia
Azoulay E et al, Chest, 2006
In 803 immunocompetent critically ill patients receiving MV for > 2 days, the
incidence of pneumonia in patients with and without respiratory-tract Candida
colonization was compared. Patients were matched on study center, admission
year, and MV duration.
%
39,7
OR 1.58
(95% CI 0.942.68)
24,1
17,6
OR 2.22
(95% CI 1.004.92)
9
VAP
4,8
Pseudomonas
VAP
colonized
8,3
extra pulmonary
candida
no-colonized
974 patients; 46 patients with candidaemia and
928 with colonisation.
Troughton JA et al. J Infect 2010;61:403-409
•A higher proportion of blood isolates were Candida
glabrata compared with non-sterile isolates (16/46 vs
106/1062; p < 0.001),
•similarly a greater proportion of blood isolates were fczresistant compared with non-sterile isolates (15/46 vs
101/1062; p < 0.001).
Troughton JA et al. J Infect 2010;61:403-409
•C. glabrata candidaemia was more likely to occur in the
absence of non-sterile site colonisation compared with
non-glabrata candidaemia (12/16 vs 8/30; p < 0.005).
•Of candidaemic patients, 43% had no preceding
colonisation by any Candida spp.; in 67% of these
patients, candidaemia was due to C. glabrata.
•Pre-emptive therapy based upon colonisation of at least
two sites may be inadequate as 43% of candidaemic
patients had no evidence of prior colonisation, 67% of
whom had candidaemia due to C. glabrata.
Troughton JA et al. J Infect 2010;61:403-409
Troughton JA et al. J Infect 2010;61:403-409
Scores and prediction rules
Prediction rules and scores for identification of nonneutropenic critically ill patients at risk for invasive
candidiasis
Candida score
The “Candida score” cut-off value is 2.5 (sensitivity 81%,
specificity 74%)
Leon C et al. Crit Care Med 2006; 34:730–737
Prediction rules and scores for identification of nonneutropenic critically ill patients at risk for invasive
candidiasis
Prediction rule
•Analysis of risk factors in 2,890 patients who stayed in the ICU for
more than 4 days
•The best prediction rule used a combination of the following factors:
-any systemic antibiotic or presence of central venous catheter and
at least two other risk factors, including
- total parenteral nutrition,
- major surgery,
- pancreatitis,
- any use of steroids and use of immunosuppressive agents.
•This prediction rule exhibited a sensitivity of 34%, a specificity of
90%, a positive predictive value of 10% and a negative predictive
value of 97%.
•This clinical rule may therefore help clinicians to rule out invasive
candididiasis.
Ostrosky-Zeichner L et al. Eur J Clin Microbiol Infect Dis 2007; 26:271-276
Prediction rules and scores for identification of nonneutropenic critically ill patients at risk for invasive
candidiasis
Is the CS useful for discriminating between Candida
colonization and invasive candidiasis in nonneutropenic critically ill patients?
•Prospective, cohort, observational study on 1107 pts
for >7 d in ICU
•A CS>=3 selected pts at high risk for IC
•IC=2.3% if CS<3
•(1–3)-Beta-D-glucan was also an independent predictor
of IC (odds ratio 1.004, 95% CI 1.0 –1.007)
Leon C et al. Crit Care Med 2009; 37:1624 –1633
Prediction rules and scores for identification of nonneutropenic critically ill patients at risk for invasive
candidiasis
Leon C et al. Crit Care Med 2009; 37:1624 –1633
Prediction rules and scores for identification of nonneutropenic critically ill patients at risk for invasive
candidiasis
Leon C et al. Crit Care Med 2009; 37:1624 –1633
Diagnosis
Diagnosing invasive fungal infections
•Given that rapid initiation of appropriate
antifungal therapy is crucial for reducing
mortality, prompt diagnosis of infection is of the
utmost importance.
•Unfortunately, diagnosing invasive fungal
infections remains difficult and is often delayed.
•Indeed, blood cultures lack sensitivity (reported
to be <50%) (Reiss E et al. Clin Microbiol Rev 1993, 6:311-323),
and usually become positive late (Morris AJ et al. J Clin
Microbiol 1996, 34:1583-1585)
Diagnosing invasive fungal infections
•Invasive tissue sampling is often problematic in critically
ill ICU patients.
•Radiological signs appear often late in the course of
infection.
•Moreover, the EORTC/MSG criteria for diagnosis of
invasive mycoses, which are based on clinical,
microbiological and radiological criteria, were developed
in immunocompromised patients and may not apply to
ICU patients.
•Need for sensitive and specific diagnostic tools nonculture based methods aimed at detecting circulating
fungal metabolites, antigens, antibodies and fungal DNA.
Mean M et al Critical Care 2008, 12:204
There are two major antigen-based tests
commercially available today:
-the mannan antigen test (Biorad) and
-the beta D-glucan test (several suppliers).
Sendid B et al. Medecine/Sciences 2009; 25:473–481.
Mannan antigen test
•The test detects the presence of mannan, a major cell
wall component of the most frequently recovered
Candida species in blood.
It is thus a Candida-specific test.
The test is intended to be used in serum or plasma.
•Because the circulation of detectable levels of
Candida antigen is of short duration in the blood, it is
recommended that frequent tests are carried out, two
to three times a week during the period at risk.
Sendid B et al. Medecine/Sciences 2009; 25:473–481.
- mannan •Antibodies to mannan usually develop when mannan
disappears; it is also recommended that antigen and
antibody detection is combined in the serial screening of
patients at risk to improve the sensitivity.
•Well designed clinical cohort studies exploring this
strategy are still rare, most of them are screening
populations of onco-hematologic patients rather than ICU
medical or surgical patients.
Verduyn Lunel FM et al. Diagn Microbiol Infect Dis 2009; 64:408–415.
Verduyn Lunel FM et al. Clin Microbiol Infect 2009; 15:380–386.
Arendrup MC et al. Clin Microbiol Infect 2009.
Ellis M et al. J Med Microbiol 2009; 58:606–615.
- mannan •The per patient sensitivity value of mannan detection
is in the range of 60%, of antimannan antibodies of
60%, and of combined tests (ag+ab) of 89%.
•The specificity of both tests was over 90%.
•A positive antigen test generally precedes positive
blood cultures or conventional diagnosis of
hepatosplenic candidiasis.
•Additional prospective cohort studies are needed to
determine the impact of mannan–antimannan detection
on the diagnosis, treatment, and outcome of invasive
candidiasis.
- mannan •The per patient sensitivity value of mannan detection
is in the range of 60%, of antimannan antibodies of
60%, and of combined tests (ag+ab) of 89%.
•The specificity of both tests was over 90%.
•A positive antigen test generally precedes positive
blood cultures or conventional diagnosis of
hepatosplenic candidiasis.
•Additional prospective cohort studies are needed to
determine the impact of mannan–antimannan detection
on the diagnosis, treatment, and outcome of invasive
candidiasis.
- beta-D-glucan •The second antigen-based test is intended to
detect the presence in plasma or serum of
beta-D-glucan (BDG), another important
component of the cell wall of the majority of
fungi, but in particular of Candida,
Aspergillus,and Pneumocystis.
•This test is thus not Candida specific.
•More than 20 studies have been published,
half as case– control, half as cohort studies.
- beta-D-glucan •Case–control studies have shown a slightly
higher sensitivity (50–90%) than cohort studies
(45–70%), and an equal specificity (70–100%) for
the diagnosis of proven or probable invasive
candidiasis.
•Two tests per week were most often performed,
and one positive test considered as positive.
•False positive tests may occur (through various
contaminated material), and the assay is
technically demanding.
Bille J et al. Curr Opin Crit Care 2010 16:460–464
β- (1,3)-D-glucan
•Given these excellent negative predictive
values β-(1,3)-D-glucan tests can help to
rule out invasive candidiasis.
•Unfortunately, little information has been
published thus far on use of β-(1,3)-Dglucan tests in the ICU setting
•A Spanish group has developed an antibody
test against the germ-tube structure of Candida
albicans cells developing when growing in
culture or invading host tissue (C. albicans IFA
IgG, Vircell, Spain).
•Applying this test to various patients
populations they have observed that patients
with positive CAGTA had a better outcome than
those with negative antibodies.
Pema´n J et al. Mycoses 2009.
Saragoza R et al. Clin Vaccine Immunol 2009; 16:1527–1528.
•Direct molecular detection of Candida DNA
circulating in blood has been tried for almost
two decades, without much success due to the
usually low yeast burden, as well as the
difficulty of extracting yeast DNA and
separating it from human DNA.
Wallet F et al. Clin Microbiol Infect 2010; 16:774–779.
•Many home-made and developed methods
have kept hope alive, with local promising
results.
•The very first commercially available PCR
method designed to detect the 25 most
prevalent microorganisms in blood culture
(comprising five species of Candida, as well as
Aspergillus fumigatus) has been evaluated for
a few years now in Europe (Septifast), and
reported in about 10 publications.
Wallet F et al. Clin Microbiol Infect 2010; 16:774–779.
SEPTIFAST
•Almost exclusively used at the time of the first blood
culture drawing and compared to the performance of
blood culture, this PCR has rarely been used as a
serial marker during the period at risk preceding the
drawing of blood culture.
•Thus it is yet not clear whether this PCR or any other
will be useful as an early marker of developing invasive
candidiasis.
The performance of the Septifast real-time multiplex PCR
10 studies, 100
febrile episodes
or samples each
SEPTIFAST
19 positive
BLOOD CULTURE
8 positive
This increased sensitivity implies mostly C. albicans,
whereas twice more C. glabrata were detected by
blood culture than by SF, but the true clinical
significance of these findings has not been
systematically assessed and reported.
Bille J et al. Curr Opin Crit Care 2010 16:460–464
NEWER TESTS
- Another commercially available multiplex PCR test
(Vyoo; SIRS-Lab, Jena, Germany) claims to detect
Candida species, but no comparative trial has been
published so far.
- An 18S rRNA broad-range PCR following an
improved DNA detection method (MolYsis; Molzym,
Bremen, Germany) has shown at least twice as many
positive results when compared to blood cultures.
Wellinghausen N et al. J Med Microbiol 2009; 58:1106–1111.
PNA FISH
Recent approaches to reduce the delay of positivity from
blood cultures:
-fluorescence in-situ hybridization test (PNA FISH
AdvanDX, Woburn, Massachusetts, USA) differentiates
the five most prevalent species of Candida.
More time consuming and expensive of FISH, allows the
identification of more species of Candida than the FISH
test does.
MALDI-TOF-MS
•The recent application to clinical microbiology of an
old technique based on the measurement of the
molecular masses of proteins and other microbial
components from whole bacterial extracts.
•This approach allows the identification of bacteria and
yeast from isolated colonies in a few minutes with an
accuracy of more than 90% when compared to
conventional identification results.
Seng P et al. Clin Infect Dis 2009; 49:543–551.
Van Veen SQ et al. J Clin Microbiol 2010; 48:900–907.
Marklein G et al. J Clin Microbiol 2009; 47:2912–2917.
Is MALDI-TOF a revolution in the microbiology
laboratory [Seng P et al. Clin Infect Dis 2009; 49:543–551] ?
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