Neonatal Candidiasis
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Transcript Neonatal Candidiasis
Neonatal
Candidiasis
Catherine M. Bendel, M.D.
University of Minnesota
Medical School,
Minneapolis
Neonatal Candidiasis Outline
Epidemiology
Risk Factors -- Yeast and Host
Research correlates
Clinical Presentation
Diagnosis and Treatment
Prophylaxis?
Candida
Commensal organism
Eucaryotic and diploid (C. albicans)
Dimorphic yeast
Increasingly important nosocomial
pathogen
Nosocomial Candidiasis
6th most common nosocomial pathogen
4th most common cause of nosocomial
septicemia (7.8% of all bloodstream infections)
2nd most common nosocomial pathogen in
NICU - incidence 2-20% in ELBW
24% to 54% mortality in those who receive
therapy
High morbidity
Pfaller. Clin Infect Dis 1996;22(Suppl 2):S89
Pittet & Wenzel. Arch Intern Med 1995;155:1177
Baltimore. Sem in Perinat 1998;22: 25
Benjamin. Pediatrics 2000;106:712
Numbers of Cases of Sepsis in the United States,
According to the Causative Organism, 19792000. Martin GL et al, N Engl J Med 348:1546-54, 2003
Virulence Factors
C. albicans accounts for the majority of
Candida spp isolated
Multiple intrinsic C. albicans virulence factors
have been identified
Systemic infections associated with specific
host risk factors
The gastrointestinal tract is considered a
major portal of entry for C. albicans
Frequency of Isolation of
Candida species
All patients
55% C. albicans
20% C. glabrata
9% C. tropicalis
7% C. krusei
15% C. parapsilosis
1% C. dubliniensis
2% others
Neonates
58% C. albicans
34% C. parapsilosis
2% C. glabrata
4% C. tropicalis
2% others
Phaller, J Clin Micro 2001:39
Fridkin, Pediatrics, 2006:117
Virulence Factors
C. albicans accounts for the majority of
Candida spp isolated
Multiple intrinsic C. albicans virulence factors
have been identified
Systemic infections associated with specific
host risk factors
The gastrointestinal tract is considered a
major portal of entry for C. albicans
C. albicans virulence factors
Adherence to host tissues (epithelium/endothelium)
Receptor or adhesin mediated
Cytolytic enzymes (proteinases, phospholipases)
Hydrophobicity
Variable phenotypic expression
Morphologic switching
Filamentous forms believed to facilitate adhesion
and penetration of epithelium
Andrutis. J Clin Microbiol 2000; 38: 2317
Cutler. Ann Rev Microbiol 1991;45:187
Mitchell. Curr Opin Microbiol 1998; 1:687
Gale. Science 1998;279:1355
Corner. Curr Biol 1997; 7:R761
Calcofluor stained cecal contents from a mouse
colonized with C. albicans showing yeast and
filamentous forms in vivo
C. albicans CAG1 (+/-) incubated 3 h on Caco-2
enterocytes showing yeast cluster partially embedded
C. albicans CAF2 (+/+) incubated 3 h on Caco-2 enterocytes with
microvilli intimately associated with distal tip of germtube
C. albicans CAF2 (+/+) incubated 3 h on Caco-2 enterocytes
showing elongated enterocyte microvilli anchoring a C.
albicans filament to the enterocyte surface
Virulence Factors
C. albicans accounts for the majority of
Candida spp isolated
Multiple intrinsic C. albicans virulence factors
have been identified
Systemic infections associated with specific
host risk factors
The gastrointestinal tract is considered a
major portal of entry for C. albicans
Host Risk Factors for Systemic Candidiasis
Colonization with Candida species
Prematurity, especially GA < 28 weeks
Very low birth weight
Prolonged hospitalization
Indwelling catheters
Neutropenia
Broad spectrum antibiotic therapy
Third generation cephalosporin exposure
Steroids
Host Risk Factors for Systemic Candidiasis
Hyperalimentation
Hyperglycemia
Indomethacin
Intestinal perforation > NEC/IP
Abdominal or cardiac surgery
Compromised anatomic barriers
Apgar score <5 at 5 min
Topical Petrolatum Ointment (Aquaphor)
NEC or IP
Strong association
IP ~25% incidence of canididasis
Empiric therapy
Incidence of Systemic Candidiasis associated
with TPO in infants with BW ≤ 1500 grams
Campbell JR, Zaccaria E, & Baker CJ, Pediatrics 2000;105:1041-1045.
Virulence Factors
C. albicans accounts for the majority of
Candida spp isolated
Multiple intrinsic C. albicans virulence factors
have been identified
Systemic infections associated with specific
host risk factors
The gastrointestinal tract is considered a
major portal of entry for C. albicans
Skin and GI tract colonization
C. albicans and
others
GI tract commensal
Acquired perinatally
Vertical transmission
from mother
Colonization within 710 days (skin & GI)
Higher levels of
colonization associated
with increased risk of
systemic disease
C. parapsilosis
Skin colonization in
adults (tinea
versicolor)
Skin or GI tract
colonization in
neonates
Acquired postnatally
Transmission from
caregivers
Colonization later,
usually >14 days: rate
higher with increased
LOS
Research
Correlates
Mouse models of IV infection and GI
tract colonization developed.
Models then used to evaluate:
The role of yeast filamentation in
colonization and infection.
The role of antibiotics and steroids in
colonization and dissemination.
Candida albicans strains
Comparisons were made using three C.
albicans strains (all URA3/ura3):
CAF-2: “wild type”/parent strain; exhibiting a
normal phenotype forming blastoconidia, germ
tubes, hyphae and pseudohyphae. (Fonzi, Genetics
1993;134:717)
HLC4: cph1/cph1,efg1/efg1; a nonfilamentous
mutant described as growing exclusively as
blastoconidida. (Lo, Cell 1997;90:939)
BCa2-10: tup1/tup1; grows exclusively in
filamentous form. (Braun, Science 1997;277:105)
Oral (107) or intravenous (105) inoculation of mice
with C. albicans CAF-2, HCL54 or BCa2-10
I.V.
C. albicans
oral
C. albicans
Antibiotics
3 days
Monitor for
death
Sacrifice 1,
7, 14, & 21
days later
C. albicans
3 days
I.P.
Dex
Sacrifice
Antibiotics in
drinking water:
• bacitracin
• streptomycin
• gentamicin
Dexamethasone:
• 2mg I.P. BID
6
6
*
4
3
-/-
-/-/+
3
*
+/+
3
-/-
4
-/-/+
4
5
-/-/+
5
*
+/+
5
Antibiotics
+ LPS
Antibiotics
-/-
No
6 antibiotics
+/+
Avg ±SE log10 viable
C. albicans /g cecum
Affect of antibiotic treatment on cecal
colonization in mice
INT1 Genotype
NOTE: C. albicans was recovered from the mesenteric lymph
nodes of only an occasional mouse and never from the
kidneys, with all treatments.
Bendel et al. SHOCK. 2000; 13:453-8.
Dexamethasone
administration
Colonization facilitated by
dexamethasone - increased to 107.5
C. albicans recovered from the
mesenteric lymph nodes in 56% of mice
C. albicans disseminated to the kidneys
in 83% of mice
Bendel et al, Pediatric Research 51:290;2002
8
Cecal Colonization
* = p<0.05
% of mice with C. albicans in tissue
Avg±SE log10 viable C. albicans /g cecum
Cecal colonization and dissemination of C. albicans to
mesenteric lymph nodes (MLN) and kidneys of orally
inoculated mice
MLN
* = p<0.05
100
100
7
6
*
5
4
3
CAF2
HLC54 BCa2-10
C.albicans strain
Kidneys
* = p<0.01
*
21/24
75
75
*
*
12/24
11/22
50
25
50
25
3/22
0/21
0/21
0
0
CAF2
HLC54 BCa2-10
C.albicans strain
CAF2
HLC54 BCa2-10
C.albicans strain
% Cumulative Survival
Mortality in mice inoculated IV with C. albicans
wild type and mutant strains
“Filamentous” only
100
80
60
“Nonfilamentous”
40
20
“Wild Type”
0
0
5
10
15
20
25
30
Days after intravenous 105 C. albicans
Avg±SE log10 viable C. albicans. per gram
Persistence of C. albicans in the kidney and liver of
intravenously inoculated mice
Kidney
8
*
*
6
*
Liver
8
* = P<.01 &
† = P<.05
vs. BCa2-10
Day 1
Day 7
Day 14
Day 21
6
*
*
4
4
†
2
2
0
0
CAF2
HLC54 BCa2-10
C. albicans strain
CAF2
HLC54 BCa2-10
C. albicans strain
A
50 microns
B
20 microns
50 microns
A
20 microns
B
PCR Results
Using specific primers for the INT1
locus of CAF-2, the EFG-A and EFG-D
fragments of HLC54, and the altered
TUP1 locus of BCa2-10, PCR was
performed on two C. albicans colonies
recovered from each cecum and one
colony from each MLN, liver and kidney
that was positive for yeast.
Results confirmed that mice were
colonized with the inoculated strain.
Summary I
All three C.albicans strains
persisted in the GI tract.
“Wild type” and “Nonfilamentous”
strains colonized in similarly high
numbers, while the numbers of
cecal “Filamentous only” present
were at least 100-fold lower.
Summary II
Of the three C.albicans strains studied:
CAF-2 (wild type) was most virulent
in IV inoculated mice as assessed by
mortality
HLC54 (defective in filamentation)
was most virulent in orally inoculated
mice as assessed by extraintestinal
dissemination of C. albicans.
BCa2-10 (constitutively filamentous)
was avirulent in both models.
Summary III
Both yeast cell and filamentous forms
of wild type and “nonfilamentous”
strains were found in kidney sections,
but tissue necrosis was associated only
with filamentous forms.
Therefore, “morphologic switching”
provides candida with an advantage for
persistence throughout the host under a
variety of conditions.
C. albicans Adherence to Cultured Epithelium - ELISA
wash, lightly fix,
wash
Incubate C. albicans with
enterocytes (pretreated for
24o with dexamethasone)
Incubate with
primary
antibody
Remove enterocytes
from plastic dish
Color
development and
O.D. at 450 nm
Secondary
antibody
conjugated to HRP
Increased adhesion of C. albicans strains to
Int 407 neonatal enterocytes following
pretreatment with dexamethasone
*
*
% Change in Adhesion
100.00%
*
100uM de x
*
1000uM de x
*
*
50.00%
*
*
*
0.00%
CAF2
10uM de x
CAG3
C. a lbi ca nsstr a in
CAG5
* p< 0.015
Increased adhesion of C. albicans strains to
Caco-2 adult enterocytes following
pretreatment with dexamethasone
100uM de x
% Change in Adhesion
100.00%
1000uM de x
*
*
50.00%
*
0.00%
CAF2
CAG3
C. a lbi ca ns str ai n
CAG5
* p< 0.05
C. albicans CAF2 (+/+) incubated 3 h on Caco-2
enterocytes showing alignment of adherent
filamentous form along enterocyte cell borders
Summary I
Dexamethasone pretreatment of enterocytes
resulted in a significant, dose dependent
increase in the adhesion of all three C.
albicans strains to both neonatal and adult
cell lines.
Neonatal enterocytes (Int407) were more
sensitive to the effects of dexamethasone
therapy. Greater increases in adhesion were
observed for all C. albicans strains, at all
concentrations, when compared to the adult
Caco-2 cell line.
Summary II
No differences were seen in the
dexamethasone effect on the adhesion of
CAG3 and CAG5, compared to CAF2 —
equivalent increases were noted to both cell
lines at each concentration tested.
Following dexamethasone treatment,
preferential adhesion of the C. albicans along
enterocyte cellular junctions was observed,
compared to more homogeneous adhesion in
the untreated cell lines.
Adhesion of Candida strains to
human epithelial cell monolayers
Clinical Candidiasis
Candida species are responsible for a wide
variety of clinical disease with variable
presentation and degree of severity
“Benign” colonization
Locally invasive disease
Catheter-related fungemia
Widespread systemic disease
The specific disease process depends on the
interplay of both host risk and yeast virulence
factors
Congenital Candidiasis - Term
Age at onset : Birth or < 24 hours
Presence of Risk Factors: None
Skin involvement: Hallmark
Respiratory involvement: Occasionally
Positive Blood culture: No
Multiorgan involvement: Never
Treatment: Topical Antifungals
Prognosis: Excellent
Congenital Candidiasis - Preterm
Age at onset : Birth or < 24 hours
Presence of Risk Factors: None
Skin involvement:
Classical candidal dermatitis
Atypical diffuse erythematous dermatitis
Respiratory involvement: Common
Positive Blood culture: Uncommon
Multiorgan involvement: Rare
Treatment: Parenteral Antifungal, avoid
central catheters
Prognosis: Guarded
Catheter-Related Fungemia
Age at onset : > 7 Days
Presence of Risk Factors: Necessary
Skin involvement: Uncommon
Respiratory involvement: Infrequent
Positive Blood culture: Yes
Multiorgan involvement: Rare
Treatment:
Parenteral Antifungal
Catheter removal
Prognosis: Good
Catheter-Related Fungemia:
Complications
Specific to site and type of catheter
Cystitis
Peritonitis
Pleuritis
Mediastinitis
Central venous catheters
Right atrial or other vascular fungal mass
Disseminated Candidiasis
Age at onset : > 7 Days
Presence of Risk Factors: Necessary
Skin involvement: None / Uncommon
Respiratory involvement: Frequent
Positive Blood culture: Yes
Multiorgan involvement: Frequent
Treatment: Parenteral Antifungal
Prognosis: Fair - Poor
Disseminated Candidiasis:
Common Complications
Pneumonia
Meningitis
< 500 WBC, lymphocytes
Normal glucose
Normal or increased protein
Renal involvement
Cystitis
Multiple or single abcesses
Pelvic fungal balls
Endophthalmitis
Significant ROP
Drohan et al, Pediatrics 2002;110(5).
Disseminated Candidiasis:
Less Common Complications
Endocarditis, pericarditis
Bone or joint involvement
Peritonitis ( with bowel perforation)
Liver abscess
Splenic abscess
Cutaneous nodules/subcutaneous
abcesses
Diagnosis
Nonspecific signs and symptoms of sepsis
Fever Uncommon
Positive Culture — NO Rapid test available
Blood
Urine (catheterized or suprapubic tap)
CSF
Other fluids as indicated
ETT, skin, mucous membrane, GI tract
cultures indicate colonization only
Diagnosis — Cultures
Fungal isolater tube not usually helpful
Cultures should be held for a minimum of 710 days: non-albicans species are slow
growing
Monitor blood cultures daily until no growth
and then weekly while on therapy
Specific Candida species should be
identified
Sensitivities to antifungal drugs should be
determined
Schelonkam RL and Moser SA, J Pediatr 2003:142:564-5
Diagnosis — Other Studies
CBC with differential and platelet count
Thrombocytopenia COMMON
WBC can be low, normal or high
Serum glucose
BUN and creatinine
Ultrasound: renal, cranial, cardiac
Ophthalmologic exam
Bilirubin and liver enzymes
Treatment — Topical
Nystatin
Miconazole
Clotrimazole
Gentian Violet
Treatment — Parenteral
Amphotericin B - mainstay of
therapy
Daily dosage:
No “test dose” required in neonates
± Initial dose 0.5 mg/kg IV over 2-6 hours
Increase by 0.5 mg/kg/d to goal of
1.0-1.5 mg/kg/d
Adjust for renal insufficiency
Treatment — Ampho B
Treatment Course
10-14 days for uncomplicated line sepsis
3- 6 weeks for disseminated or complicated
sepsis. Cumulative dose of 30-35 mg/kg or
clearance of disease — whichever comes
first!
Monitor systemic involvement for
improvement/clearance — serial ultrasounds,
repeat cx, etc.
Treatment — Ampho B
Complications of therapy:
Renal insufficiency
Monitor UOP, BUN,Cr qod initially; then q week if stable
Renal failure reversible, but dialysis may be required
Profound hypokalemia / hypomagnesemia
Monitor K and Mg levels closely
Hematologic - bone marrow suppression
Monitor CBC and platelets qod initially and then q week
Liver dysfunction
Adjunctive Therapy: 5-FC
5-fluorocytosine - Synergistic
Dosing:
50-100 mg/kg/day po divided q 12 hours
Monitor levels
Limitations:
Oral
Rapid resistance occurs if used alone
Alternative Therapy
Liposomal Amphotericin B (Ambisome) or
Amphotericin B Lipid Complex (Ablecet) or
Amphotericin B colloidal dispersion
Dosing: 3-5 mg/kg/day IV over 2hours
Appears to be safe and effective, but not
superior to conventional Ampho B
Diminished side effects, especially renal
Limitations:
Decreased renal absorption
Comparison of Antifungal medications
Kicklighter SD. NeoReviews 3;12:249-255. 2002
Medication
Route Daily Dose Cost/500 mg
(mg/kg/24) (acquisition)
Amphotericin B
IV
Fluconazole
IV,PO 3 - 6
$66
Abelcet
IV
3-4
$93
Amph B Colloid Disp IV
5
$135
Ambisome
IV
3-5
$199
Flucytosine
PO
50 - 200
$7
0.5 - 1.5
$12
Alternative Therapy
Fluconazole — IV (vorconazole next generation)
Dosing, over 2-6 hours:
Preterm ≤ 29 weeks: 5-6 mg/kg/72 hours
Preterm 30-36 weeks: 3-6 mg/kg/48 hours
Term: 6-12 mg/kg/24-72 hours
Monitor levels
Side effects: renal, hepatic and
hematologic, but less than Ampho B
Yeast Susceptibilities
Fairview-University Medical Center – 2009
Candida
albicans
Candida
glabrata
Candida
krusei
Candida Candida
tropicalis parapsilosis
Ampho B
99% 99% 100% 100% 100%
5-FC
99% 98%
0%
93% 99%
Fluco nazole
97% 44% O%
79% 95%
Itraco nazole
92% 26% 39% 77% 95%
Alternative Therapy
Echinocandin
Caspofungin
Micafungin
? Prophylaxis ?
Key to preventing disease appears
to be diminishing colonization and
risk of dissemination.
Chemoprophylaxis ?
Nystatin / Mycostatin (?efficacy, benign)
Fluconazole (?efficacy, ? benign)
Molecularly generated inhibitory proteins?
Fluconazole Prophylaxis
Kaufman et al, NEJM 200;345
IV fluconazole for the first six weeks of life in
infants with BW ≤ 1000g
Decreased levels of colonization (20 v 60%)
Fewer invasive fungal infections (0 v 20%)
Safe, no change in sensitivity patterns
Caveats
Extremely high rate of invasive infections
TPO/Aquaphor used
Unable to determine pressure for resistance
.
Many thanks to all who
have helped with the
research!
Adhesion studies: Mary
Keane & Beth Swanson
Animal studies: MaryAlice Johnson & Robb
Garni
Microscopy: Stanley
Erlandson & Michelle
Henry-Stanley
Everything: Carol Wells