Cryptococcosis John R. Perfect, MD a, * , Arturo Casadevall, MD, PhD b a Department of Medicine, Division of Infectious Diseases, Duke University Medical Center, PO Box 3353, Durham, NC 27710, USA b Department of Medicine, Division of Infectious Diseases, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461, USA Epidemiology Varieties of C. neoformans Cryptococcus neoformans strains classically have been grouped into two varieties that included five serotypes known as C. neoformans variety neofor- mans (serotypes A, D, and AD) and C. neoformans variety gattii (serotypes B and C). The serotype differences between the different varieties reflect anti- genic differences resulting from differences in the structure of the capsular polysaccharide. The serotype classification of a strain is determined using absorbed rabbit sera [1]. Some monoclonal antibodies are now available that are useful for typing strains [2,3]. C. neoformans var. neoformans is found throughout the world in association with excreta from certain birds including pigeons, canaries, and cockatoos. C. neoformans var. gattii is found primarily in tropical and subtropical regions and has been associated with several species of eucalyptus trees. After many years of stable taxonomic classification the availability of DNA typing data has shown major differences between isolates grouped within a serotype and, as a consequence, there is now considerable uncer- tainty as to the exact relationship between the varieties and serotypes. The situation is made more complex by the fact that C. neoformans has a sexual cycle and that some strains are capable of mating with consequent genetic recombination. Recently it was proposed to separate serotype A into a different variety known as grubii on account of significant genetic dif- ferences from serotype D [4]. The varietal status for the serotype AD strains Infect Dis Clin N Am 16 (2002) 837–874 * Corresponding author. E-mail address: [email protected] (J.R. Perfect). The authors are supported by the National Institutes of Health with NIAID grants AI33774, AI3342, and HL59842 (AC); and AI28388, AI49975, PA 98100, and AI39115 (JRP). 0891-5520/02/$ - see front matter Ó 2002, Elsevier Science (USA). All rights reserved. PII: S 0 8 9 1 - 5 5 2 0 ( 0 2 ) 0 0 0 3 6 - 3
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Cryptococcosis
John R. Perfect, MDa,*, Arturo Casadevall, MD, PhDb
aDepartment of Medicine, Division of Infectious Diseases,
Duke University Medical Center, PO Box 3353, Durham, NC 27710, USAbDepartment of Medicine, Division of Infectious Diseases, Albert Einstein
College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461, USA
Epidemiology
Varieties of C. neoformans
Cryptococcus neoformans strains classically have been grouped into two
varieties that included five serotypes known as C. neoformans variety neofor-
mans (serotypes A, D, and AD) and C. neoformans variety gattii (serotypes
B and C). The serotype differences between the different varieties reflect anti-genic differences resulting from differences in the structure of the capsular
polysaccharide. The serotype classification of a strain is determined using
absorbed rabbit sera [1]. Some monoclonal antibodies are now available
that are useful for typing strains [2,3]. C. neoformans var. neoformans is
found throughout the world in association with excreta from certain birds
including pigeons, canaries, and cockatoos. C. neoformans var. gattii is
found primarily in tropical and subtropical regions and has been associated
with several species of eucalyptus trees.After many years of stable taxonomic classification the availability of
DNA typing data has shown major differences between isolates grouped
within a serotype and, as a consequence, there is now considerable uncer-
tainty as to the exact relationship between the varieties and serotypes. The
situation is made more complex by the fact that C. neoformans has a sexual
cycle and that some strains are capable of mating with consequent genetic
recombination. Recently it was proposed to separate serotype A into a
different variety known as grubii on account of significant genetic dif-ferences from serotype D [4]. The varietal status for the serotype AD strains
The authors are supported by the National Institutes of Health with NIAID grants
AI33774, AI3342, and HL59842 (AC); and AI28388, AI49975, PA 98100, and AI39115 (JRP).
0891-5520/02/$ - see front matter � 2002, Elsevier Science (USA). All rights reserved.
PII: S 0 8 9 1 - 5 5 2 0 ( 0 2 ) 0 0 0 3 6 - 3
is uncertain but there is increasing evidence that these represent diploid
strains, possibly resulting from matings of A and D strains. There is now
a further proposal to abolish the current varietal system and replace it withtwo species that contain groupings of genotypes [5]. Given the impending
completion of the C. neoformans genome and rapid advances in the under-
standing of genetic diversity among fungi it is likely that taxonomic relation-
ship of strains will remain in flux for some time. For clinicians, however, the
standard serotype classification, which has been in use for half a century,
remains a useful method for approaching strain differences.
Most clinical isolates are serotype A (variety grubii) even in geographic
regions where strains of variety gattii are found in the environment [6,7].Serotype D is common in certain European countries but constitutes a
minority of isolates in other parts of the world [6,7]. For example, in New
York City serotype D comprises approximately 13% of isolates [8]. From
a clinical viewpoint there has been little emphasis in determining the sero-
type of clinical strains because all varieties produce similar forms of disease
and there do not seem to be significant differences in drug susceptibility
among the various isolates. Hence, strain serotyping is not routinely per-
formed by microbiology laboratories and serotype information is seldomavailable to clinicians. There is increasing evidence, however, that the differ-
ent varieties result in different clinical manifestations or clinical syndromes.
For example, cryptococcosis caused by C. neoformans var. gattii occurs
primarily in immunologically competent hosts and is associated with the
formation of large granulomas (cryptococcomas) that may require surgical
removal for optimal management [9]. Serotype D strains have been associ-
ated with a propensity to cause cutaneous lesions [10].
Pathogenesis and prevalence of human C. neoformans infection
In the 1950s, Baker et al [11–13] carried out autopsy studies and docu-
mented the existence of a primary infection complex in individuals with and
without a history of cryptococcosis. These studies established that the
pathogenesis of cryptococcosis was similar to that of Mycobacterium tuber-
culosis and involved an initial pulmonary infection that was contained with-
in the lung by granulomatous inflammation. Evidence that latency canfollow an initial infection and reactivate later in life comes from DNA typ-
ing studies of C. neoformans isolates from African expatriates who devel-
oped cryptococcosis while living in Europe [14]. In this study isolates
from African expatriates were more similar to those from Africa than to the
prevailing European strains [14]. Other evidence consistent with the exis-
tence of a latent state is the description of cryptococcosis caused by C. neo-
formans var. gattii strains in individuals living outside tropical areas with a
history of travel [15]. There is also evidence, however, that initial exposuresto the yeast can lead to symptomatic infection. For example, there are anec-
dotal reports of cryptococcosis following direct exposure to pigeons and
838 J.R. Perfect, A. Casadevall / Infect Dis Clin N Am 16 (2002) 837–874
their excreta [16]. Recently, an immunocompromised patient was shown to
have cryptococcosis with a strain that was indistinguishable from an isolate
found in the excreta of a pet bird in the home [17]. Hence, initial acquisitionof C. neoformans from the environment can presumably lead to infection
with at least three outcomes: clearance of infection, development of latent
infection, or acute infection with or without disseminated cryptococcosis.
The outcome of infection is presumably a function of the inoculum, the
immunologic state of the host, and the virulence of the infecting strain.
At this time the interaction of these factors to determine precisely the out-
come of infection remains poorly understood but it is believed that most pri-
mary C. neoformans infections are asymptomatic.The prevalence of infection with C. neoformans is believed to be high even
when symptomatic disease is rare. Several methods have been used to study
the prevalence of infection with C. neoformans among individuals without
evidence of cryptococcosis. Skin testing for delayed hypersensitivity to C.
neoformans antigens has shown positive responses in individuals with a his-
tory of known exposure to the fungus [18]. The prevalence of positive skin
reactions in individuals with presumed high exposure to C. neoformans, such
as pigeon fanciers and laboratory workers in cryptococcal research labora-tories, is high suggesting that asymptomatic infection is common [19]. Sero-
logic studies have consistently provided evidence for widespread infection in
individuals without a history of cryptococcosis. Practically all adults have
serum antibody to C. neoformans antigens [20]. Analysis of sera from chil-
dren of different ages in New York City has shown that the age of serocon-
version for most individuals is before the age of 10 [21]. Interestingly, that
study reported one child who presented to the emergency room with vomit-
ing and fever and was positive for serum cryptococcal antigen, which raisesthe issue that some childhood infections are accompanied by antigenemia
and may be an early indicator of disseminated infection [21].
Although there are reports of occasional isolation of C. neoformans from
individuals with no signs or symptoms of cryptococcosis [22,23], this fungus
is not generally considered to be a normal constituent of the human micro-
bial flora. The likelihood of recovering C. neoformans from asymptomatic
humans increases, however, in individuals with chronic pulmonary diseases
[24,25]. Hence, isolation of C. neoformans from a human specimen shouldprompt consideration of the possibility of chronic infection.
Incidence on human cryptococcosis
Despite strong serologic evidence for widespread infection in certain
human populations, cryptococcosis is a rare disease in individuals without
impaired immunity. Before the AIDS epidemic, cryptococcosis was consid-
ered an unusual infection and there was a dearth of robust epidemiologicstudies to provide the true incidence and prevalence of cryptococcosis in the
absence of HIV infection. A retrospective review of the Kaiser-Permanente
839J.R. Perfect, A. Casadevall / Infect Dis Clin N Am 16 (2002) 837–874
records in Northern California for the years 1971 to 1980 in the pre-AIDS
era yielded only 10 cases with an overall incidence of 0.8 cases per million
persons per year [26]. Extrapolation of these numbers to calculate a rate forthe United States per year would have yielded 176 cases [26], which is not far
off from the estimates of 200 to 300 cases per year made in the 1960s [27]. A
retrospective analysis of the data at the Centers for Disease Control and
Prevention, however, revealed a steady rise in the number of identified cases
through serologic testing from 24 cases in 1965 to 336 in 1976 [27]. Whether
these numbers reflected increased awareness of the infection, improvements
in serologic detection, or a true increase in the incidence of cryptococcosis
cannot be discerned from the information available. Nevertheless, it is clearthat in the 1970s there was an increasing recognition of cryptococcal disease,
which may have reflected the beginning of the AIDS epidemic, the increas-
ing use of organ transplantation to treat certain incurable diseases, and
progress in cancer therapy resulting in larger numbers of immune-sup-
pressed survivors. A recent population-based study using active surveillance
has documented that the incidence of cryptococcosis in non–HIV-infected
individuals varies with geographic location and ranges from 0.2 to 0.9 per
100,000 in Atlanta and San Francisco, respectively [28]. An active surveil-lance study in Alabama found an overall annual incidence of 0.84 cases per
100,000 of cryptococcosis in non–HIV-infected individuals [29]. Rates of
cryptococcosis approaching 1 case per 100,000 in patients without HIV
infection are comparable with the incidence rates described for meningococ-
cal meningitis [30].
After the recognition of the AIDS epidemic in 1981 cryptococcosis in
individuals without an obvious cause of immune impairment became an
AIDS-defining diagnosis. Many studies documented a high prevalence ofcryptococcosis in AIDS patients with rates ranging from 2.9% to 13.3%
[31–37]. The annual incidence of cryptococcosis in patients with advanced
HIV infection in 1992 to 1994 was 17 and 66 per 1000 persons living with
AIDS in San Francisco and Atlanta, respectively [28]. These incidence num-
bers predate the introduction of more effective antiretroviral therapy, which
has been associated with a decline in the prevalence of cryptococcosis in this
population (see later).
Two other groups are at significantly higher risk for cryptococcosis: can-cer patients and recipients of organ transplants. Lymphoproliferative dis-
orders were associated with an increased risk for cryptococcosis in the
1950s [38,39]. Subsequent studies have confirmed a continuing association
between cryptococcosis and hematologic malignancies [40–42]. Although
prospective population-based incidence data for cryptococcosis in patients
with cancer are not available a retrospective analysis of case records at the
M.D. Anderson Cancer Center (Houston, TX) for the years 1989 to 1999
revealed an incidence of 18 cases per 100,000 admissions. Cryptococcosisoccurred in 2.8% of organ transplant recipients with an overall associated
mortality of 42% [43]. Kidney and liver transplant recipients seemed to be
840 J.R. Perfect, A. Casadevall / Infect Dis Clin N Am 16 (2002) 837–874
at high risk for cryptococcosis and there was a significantly higher preva-
lence of C. neoformans infection among organ transplant recipients in the
Northeastern region of the United States [43]. Furthermore, both in cornealand lung transplantation, the transplanted organ has carried the cryptococ-
cal infection to the host [44,45].
Sources of infection
Cryptococcus neoformans var. gattii (serotype B) is found in several spe-
cies of eucalyptus trees, whereas C. neoformans varieties neoformans (sero-
type D) and grubii (serotype A) have been isolated from various sourcesincluding fruits, trees, and avian excreta [46]. There is general agreement
in the field that most cryptococcal infections are acquired by inhalation of
infectious propagules. The specific source of infection and the form of C.
neoformans responsible for human infection, however, have not been defin-
itively established. Encapsulated yeast cells of the type found in laboratory
cultures and in clinical isolates are thought to be too large to be inhaled effi-
ciently. Dehydrated yeast cells or basidiospores (<5 to 10 lm), however, are
sufficiently small for inhalation and alveolar deposition. Analysis of soilscontaminated with pigeon excreta has shown the presence of large numbers
of propagules with diameters compatible with alveolar deposition [47,48].
Air in sites heavily contaminated with pigeon excreta has also been shown
to contain viable cryptococci. Analysis of the air in a tower in Oklahoma
City contaminated with pigeon excreta revealed an average concentration
of 45 viable cells of C. neoformans cells per 100 L of air with approximately
60% of the cells being less than 5 lm in diameter [49]. Based on these num-
bers it was estimated that a human exposed to this air for 1 hour would have41 C. neoformans cells deposited in the lungs [49]. Several studies have
shown that clinical isolates are indistinguishable by molecular typing from
environmental isolates [50–52].
Direct evidence for human infection from either eucalyptus trees or
pigeon excreta is not yet available. The finding of viable C. neoformans cells
of size compatible with alveolar deposition in air from sites contaminated
with pigeon excreta and the association of DNA types in clinical- and
excreta-derived isolates, however, provides strong circumstantial evidenceimplicating this material as a potential source of infection. There is one
well-documented case that suggests acquisition of C. neoformans infection
from a pet cockatoo [17]. It is advisable that individuals at risk for crypto-
coccosis avoid sites that are contaminated with pigeon excreta and desist
from having certain birds, such as pigeons, canaries, and cockatoos, as pets.
Although there is not a strong seasonal association for the occurrence of
cryptococcosis, some studies have noted a trend toward higher rates in the
fall and winter [37]. A study from Thailand, however, noted no seasonalassociation for C. neoformans infections while demonstrating a strong asso-
ciation for Penicillium marneffei infection in the dry season [53].
841J.R. Perfect, A. Casadevall / Infect Dis Clin N Am 16 (2002) 837–874
Risk factors
Life-threatening cryptococcal disease can occur in apparently normal
individuals but such cases are relatively rare. The likelihood of cryptococco-sis rises dramatically, however, in individuals with impaired immunity. Spe-
cifically, conditions that predispose to impaired cellular immunity have been
associated with a significantly increased risk of cryptococcosis and these
include advanced HIV infection [32], lymphoproliferative disorders [39], ste-
roid therapy [54], and organ transplantation [55]. Cryptococcosis has also
been associated with hyper-IgM syndrome in children [56–58] and HIV-
negative CD4+ T lymphopenia in adults [59]. A smoking habit may predis-
pose to C. neoformans infection in certain individuals [28,60]. Population-based studies have also yielded an association between outdoor activities
and the risk of acquiring cryptococcosis [60].
In patients with HIV infection the incidence of cryptococcosis is inversely
proportional to the CD4 lymphocyte count [61]. In patients with HIV infec-
tion the risk of cryptococcosis rises rapidly when the CD4 lymphocyte count
drops below 100/lL [37]. The average CD4 lymphocyte count of HIV-
infected patients with cryptococcosis has been measured at 73/lL [61].
Current trends in the epidemiology of cryptococcosis
After rising dramatically until the early 1990s, the prevalence of crypto-
coccosis in patients with HIV infection began to decline with the introduc-
tion of more effective antiretroviral therapy and the widespread use of
fluconazole therapy for the treatment of oral candidiasis [28]. Temporal
trends noted a rising incidence of cryptococcosis in HIV-infected patientsfrom 1985 to 1992 [62], but by the mid-1990s there was evidence for fewer
infections in this population [63]. The introduction of highly active anti-
retroviral therapy (HAART) reduces viremia and increases CD4 counts,
resulting in an improved immunologic state that is associated with signifi-
cantly reduced risk for cryptococcosis. In The Netherlands the number of
patients with HIV infection complicated by C. neoformans infection has
declined by as much as 75% with the introduction of HARRT [64]. Another
important contributor is probably the use of fluconazole for the suppressionof oropharyngeal candidiasis in patients with AIDS [65]. In this regard flu-
conazole therapy has been shown to be effective prophylaxis against the
development of cryptococcal meningitis [66,67].
Because HIV infection is currently the major risk factor for cryptococco-
sis in countries affected by the AIDS epidemic the use of HAART has been
associated with a reduced incidence of cryptococcosis in areas where it is
available. The high cost of HAART and the complicated logistical infra-
structure that is necessary for its administration, however, have precludedits use in less-developed countries of Africa and Asia where the prevalence
of HIV infection and AIDS increased dramatically in the 1990s and so has
842 J.R. Perfect, A. Casadevall / Infect Dis Clin N Am 16 (2002) 837–874
cryptococcosis. For example, in Asia a prospective study of bloodstream
infections in patients hospitalized at a Bangkok hospital revealed that C.
neoformans was the most common pathogen recovered in culture [68]. Thenet result of these epidemiologic trends has been an explosion in the inci-
dence of cryptococcosis in Africa, Thailand [68], and India [69], whereas the
number of cases in Europe and the United States has declined.
Several reports early in the AIDS epidemic suggested that cryptococcosis
was more prevalent in Africa than in other geographic regions [70,71]. Cur-
rently, cryptococcal meningitis is the leading cause of culture-positive men-
ingitis in Zimbabwe, constituting 45% of all cases and easily outnumbering
the cases of pyogenic (16%) or tuberculous (12%) meningitis [72]. A similarsituation exists in Malawi where C. neoformans is the most common cause of
meningitis having surpassed both Neisseria meningitidis and Streptococcus
pneumoniae in this role [69]. The mortality in African patients is high, aver-
aging over 40%, and the mean duration of symptoms before hospitalization
is only 4.8 days [69]. The exact reason for the apparently higher prevalence
of cryptococcosis in African patients with HIV infection is not known.
There is some evidence that the incidence of cryptococcosis in Africa has
been increasing for decades possibly as a consequence of the developingHIV epidemic in that continent [73]. Interestingly, a population analysis
of AIDS-defining illnesses in Australia revealed that birth in Africa was
associated with an odds ratio for developing cryptococcal meningitis that
was 2.43 times that observed for individuals born in Australia or other
industrialized nations [74]. The authors speculate that the higher likelihood
of cryptococcal disease in African expatriates may be a result of greater
exposure and lower rates of antifungal prophylaxis [74]. It is also possible,
however, that host genetic and nutritional differences, possibly combinedwith C. neoformans strain differences, are responsible for the greater preva-
lence of cryptococcosis in sub-Saharan patients with AIDS.
Host defense and immune response
There is evidence that C. neoformans infections are common in the gen-
eral population based on serologic [21] and skin hypersensitivity [75,76]
studies. Because the prevalence of infection in the general population seemsto be very high yet the incidence of cryptococcosis is relatively low, one must
conclude that normal host defense mechanisms are highly effective at con-
taining and preventing disease. Nevertheless, the reality of reactivation dis-
ease in many cases of human cryptococcosis [14] strongly suggests that the
establishment of latent infection in granulomas may be a relatively common
event following acute infection in normal individuals. C. neoformans
presents the paradox that normal immune mechanisms must be highly effec-
tive at containing infection but may be unable to eradicate the organismonce established in tissue in many immunocompetent hosts. The immune
response to C. neoformans has been studied extensively in humans and
843J.R. Perfect, A. Casadevall / Infect Dis Clin N Am 16 (2002) 837–874
experimental animals and is relatively well understood. The mechanisms by
which C. neoformans subverts the immune system to establish chronic infec-
tions are only beginning to be unraveled.
Effective immune response
Most cryptococcal infections are associated with conditions whereby cell-
mediated immunity is compromised [77]. This observation strongly suggests
an important requirement of cell-mediated immunity in host defense against
C. neoformans and there is general agreement that a strong cellular immune
response producing granulomatous inflammation is essential for contain-ment of C. neoformans infection [78–81]. In humans, containment of crypto-
coccal infection in the lung is associated with the formation of a primary
complex, which consists of a granulomatous subpleural nodule [11]. In brain
tissue a granulomatous response is associated with control of infection in
both humans [82] and rats [82,83]. Experimental cryptococcal infection in
immunocompetent rats reveals that control of infection with diminution
of fungal burden is temporally associated with a granulomatous tissue reac-
tion [78]. Similarly, normal rabbits infected with C. neoformans mount astrong cellular immune response in the meninges that is associated with con-
tainment of an intracisternal infection [84]. Because granuloma formation is
a result of a Th1-polarized response, there is a requirement for the produc-
tion of Th1-associated cytokines, such as interferon-c and interleukin (IL)-2,
and effective host responses to C. neoformans infection [85,86].
The macrophage is generally considered to be the central effector cell
against C. neoformans [87–89]. Alveolar macrophages are probably the first
immune effector cell encountered by inhaled cryptococcal cells [90,91]. Themacrophage functions in defense against cryptococcosis by ingesting and
killing yeast cells and by producing proinflammatory cytokines, such as
IL-12. Chemokines, such as MCP-1 and MIP-1a, are important for recruit-
ment of inflammatory cells to sites of cryptococcal infection [92]. Histologic
examination of infected tissue reveals that macrophages are closely associ-
ated with cryptococci [87]. T cells are believed to contribute to host defense
by providing cytokines that activate macrophage fungicidal activity, such as
interferon-c [93], and by promoting the transformation of alveolar macro-phages into giant cells capable of ingesting large encapsulated yeast cells
[94]. Natural killer cells, neutrophils, eosinophils, and certain types of lym-
phocytes have each been reported to mediate direct antifungal effects in vitro
suggesting that they contribute to host defense [46].
There is evidence that humoral immunity contributes to protection
against C. neoformans [95]. Four independent research groups have shown
that passive administration of certain monoclonal antibodies to the polysac-
charide capsule can prolong survival or reduce fungal burden [96–99]. A pol-ysaccharide-tetanus toxoid conjugate vaccine was shown to elicit high titers
of antibody to the capsule that protected against an intravenous infection in
844 J.R. Perfect, A. Casadevall / Infect Dis Clin N Am 16 (2002) 837–874
mice [100]. The availability of specific antibody can help the host response
by providing opsonins for efficient phagocytosis, enhancing natural killer
cell function, and clearing capsular polysaccharide [101]. Antibodies to glu-cosylceramide [102] and melanin [103] in the cell wall have been shown to
modify the course of infection suggesting the existence of other targets for
humoral immunity in addition to the polysaccharide capsule. Serologic stud-
ies in patients at risk for infection suggest qualitative and quantitative differ-
ences in the types of immunoglobulins present that may predispose to
disseminated disease [104].
Intracellular pathogenesis
Histologic examination of C. neoformans–infected tissue can reveal
diverse inflammatory responses ranging from the virtual absence of tissue
reaction to intense granulomatous inflammation resembling caseous
necrosis [79]. The budding index of C. neoformans in tissue is inversely pro-
portional to the intensity of the granulomatous reaction which is consistent
with the view that strong cell-mediated tissue responses are required to con-
tain infection [79]. The protean nature of tissue responses to this fungusseems to be a function of both the immunologic status of the host and cer-
tain yet undefined characteristics of C. neoformans. For example, switch var-
iants of a single C. neoformans strain can elicit diverse inflammatory
responses in rats ranging from minimal inflammation to caseous necrosis
[105]. There is evidence that the size of the capsule affects the type of
response. In this regard, infection with small capsule variants often elicits
intense inflammatory responses that have been confused with Histoplasma
capsulatum [106]. This phenomenon is believed to represent interference ofcapsular polysaccharide during infection with these well-encapsulated
strains and the inflammatory response (see later).
The location of C. neoformans vis-a-vis inflammatory cells is also a func-
tion of the inflammatory response. Although the capsular polysaccharide
is antiphagocytic in vitro in conditions where there are no complement or
antibody opsonins, the fungus is rapidly ingested by alveolar macrophages
after experimental infection [107]. Granulomatous responses are associated
with a predominance of intracellular forms, whereas in the absence of in-flammation cryptococci can grow as extracellular fungal masses, which are
grossly visible in tissue and can give a ‘‘soap suds’’ appearance [79]. There is
increasing evidence, however, that C. neoformans can survive in macro-
phages for prolonged periods of time [89]. Studies on the course of C. neo-
formans infection in immunocompetent mice have shown that C. neoformans
can replicate inside alveolar macrophages resulting in host cell lysis [107].
At the initial stages of infection, before the immune response is established,
most of the replication occurs in the intracellular compartment. C. neofor-mans has a unique strategy of intracellular replication in macrophages
whereby capsular polysaccharide is secreted into vesicles resulting in
845J.R. Perfect, A. Casadevall / Infect Dis Clin N Am 16 (2002) 837–874
disruption of the vesicular network, which presumably leads to cyto-
toxicity [108].
From a clinical viewpoint the occurrence of intracellular parasitism in C.
neoformans infection is significant because it suggests that efficient eradica-
tion of infection requires antifungal agents capable of host cell penetration
and that are active in phagolysosomes.
Immunomodulation by C. neoformans and its products
There is considerable evidence that C. neoformans infection is able to
modulate the host immune response through various fungal products. C.
neoformans infection elicits suppressor responses in experimental animals,which presumably down-regulate the cellular immune response to infection
[109,110]. In rats, the maintenance of chronic infection is associated with
down-regulation of both humoral and cellular immune responses [89]. The
best characterized immunomodulator is the capsular polysaccharide, which
has been shown to interfere with a variety of immunologic functions [111].
C. neoformans sheds capsular polysaccharide into tissue during infection,
which presumably results in both local and global inhibition of the immune
response [111]. Locally, shed polysaccharide may interfere with leukocytemigration, cytokine production, and phagocytosis, whereas globally this
antigen is believed to elicit a state of immunologic unresponsiveness such
that affected hosts seldom mount significant antibody responses [112].
In addition to exopolysaccharides, C. neoformans synthesizes three other
products that can affect the immune response: (1) mannitol, (2) melanin, and
(3) prostaglandins. C. neoformans produces mannitol in tissue [113], which
can interfere with oxidative killing by phagocytic cells [114]. C. neoformans
has a laccase that can synthesize melanin from phenolic precursors [115].Melanin production in tissue has been documented during both human and
rodent experimental infection [116,117]. Melanin is believed to contribute to
virulence by protecting the fungal cell against oxidative and nonoxidative
fungicidal mechanisms of host immune effector cells [118]. Melanin may also
act as an immunomodulator by down-regulating the cellular immune
response in the lung [119]. C. neoformans has recently been shown to pro-
duce prostaglandins in vitro that could modulate the immune response if
they are produced in vivo [120].
Clinical manifestations
There are two primary sites of infection with C. neoformans: the lung and
the central nervous system (CNS). In a recent review of non–HIV-infected
patients, there were 109 (36%) with only pulmonary involvement and 157
(51%) with CNS disease and the rest with other or multiple sites [121]. Threeother sites of infection (the skin, prostate, and eye) have particular clini-
cal features that require some specific clinical attention. It should be
846 J.R. Perfect, A. Casadevall / Infect Dis Clin N Am 16 (2002) 837–874
emphasized, however, that C. neoformans can infect any organ in the body
[122]. There are also a few differences that distinguish cryptococcosis in
patients with or without HIV infection, such as more CNS and extrapulmo-nary involvement, higher rate of positive India ink examinations, positive
blood cultures, and fewer cerebrospinal fluid (CSF) inflammatory cells. This
is primarily a distinction of severity of immunosuppression and burden of
yeast and not a specific feature or tropism between HIV and the fungus.
Lung
This organ is the most common portal of entry for infection and symp-toms range from asymptomatic colonization [123] to life-threatening pneu-
monias [124]. Although at least a third of normal hosts are asymptomatic
and infection is found by an abnormal chest radiograph, patients may
present with evidence of acute infection, such as fever, chest pain, cough,
weight loss, and sputum production [125,126]. There are also a series of
unusual presentations of pulmonary cryptococcus, such as ‘‘allergic’’ cryp-
obliterans with organizing pneumonia [129], and superior vena cava syn-drome [130,131]. It is also possible to have simultaneous infection with
other pathogens, such as tuberculosis, Nocardia, and Echinococcus [132–135].
The chest radiographs in apparently normal hosts range from well-defined,
noncalcified, single or multiple lung nodules; indistinct mass-like infiltrates;
hilar lymphadenopathy; pleural effusions; and lung cavitation [136]. There is
no characteristic radiograph, although single or multiple nodules are most
common and may radiographically mimic a malignant tumor. If the infec-
tion is limited to the lung, the serum cryptococcal antigen is generally neg-ative and in the authors’ opinion a positive cryptococcal serum antigen is a
sign that the yeast may have disseminated from the lung to other body sites.
Patients with C. neoformans isolated from the lung and with an underlying
immunosuppressive risk factor for invasive infection should probably be
considered for lumbar puncture to rule out simultaneous CNS infection
with or without symptoms. In the apparently normal, asymptomatic patient
with C. neoformans isolated from the lung, however, the yield of positive
findings with lumbar puncture is so low that it does not need to be per-formed routinely.
In the severely immunocompromised host, cryptococcal pneumonia can
progress more rapidly; the ability to disseminate outside the primary lung
focus to the CNS allows patients to present with a meningeal rather than
a pulmonary syndrome [137]. Patients can, however, develop an over
whelming cryptococcal pneumonia with features of adult respiratory distress
syndrome [138–140] without CNS involvement. Unlike immunocom-
petent patients, most immunosuppressed patients have constitutionalsymptoms, such as fever, malaise, chest pain, cough, dyspnea, weight
loss, or headaches. Chest radiographs are similar in spectrum to
847J.R. Perfect, A. Casadevall / Infect Dis Clin N Am 16 (2002) 837–874
nonimmunocompromised hosts except that alveolar and interstitial infil-
trates may be more common and can potentially be confused with other
pathogens, such as Pneumocystis carinii. In AIDS patients, cryptococcalpneumonia is primarily symptomatic and over 90% already have concomi-
tant CNS infection at clinical presentation [124]. This observation reflects
the extreme nature of the immunodeficiency in these patients at the time
of cryptococcosis in which CD4 counts have generally fallen under 100
cells/lL. It should also be noted that all severely immunocompromised
patients with pulmonary cryptococcosis may present with other concomi-
tant opportunistic pulmonary and disseminated infections, such as
typical or atypical mycobacteria, cytomegalovirus, Nocardia, and P. carinii
infections.
Central nervous system
Most patients with cryptococcal meningitis present with signs and symp-
toms of subacute meningitis or meningoencephalitis, such as headaches,
fever, lethargy, coma, or memory loss over 2 to 4 weeks. Patients may not
have classic symptoms, however, and present with acute (several days)symptoms of severe headaches, intermittent headaches, or no headaches
with altered mental status. It is difficult to separate AIDS patients from
others in their presentation except to point out that symptoms may be
shorter; there is a greater chance of finding a secondary site of infection; a
greater likelihood of having a second CNS event with either infections, such
as Toxoplasma gondii, or lymphoma; and a higher burden of yeasts. A recent
new cryptococcal syndrome associated with HAART and partial immune
reconstitution has been described during HIV infection. Several patientsafter starting or changing HAART developed acute symptoms of cryp-
tococcal meningitis or pain and swelling in tissues [141]. It is hypothesized
that with an improved immune system produced by HAART, a silent or
latent cryptococcal infection was then clinically made apparent as an inflam-
matory reaction was mobilized to the clinically silent yeast cells or polysac-
charide antigen [141].
There are little data that relate the severity of CNS disease to the infecting
strain of C. neoformans and it is generally considered that the state ofhost defenses primarily determines clinical manifestations. There are some
suggestions, however, that certain strains can influence clinical presen-
tation. For instance, in Australia, which attends to infections with both
C. neoformans var. neoformans-grubii and C. neoformans var. gattii, cerebral
cryptococcomas or hydrocephalus with pulmonary mass lesions in immuno-
competent host, were generally found with the variety gattii infections [142].
In general, infection with this variety has longer survival rates than those
with variety neoformans-grubii. In a subgroup of patients with variety gattii
infection, however, there are more brain-enhancing lesions by scan, compli-
cations of hydrocephalus, cranial nerve palsies, and elevated intracranial
848 J.R. Perfect, A. Casadevall / Infect Dis Clin N Am 16 (2002) 837–874
pressures, and a poor prognosis is noted [142,143]. Although these findings
may relate to the distribution of infection in normal hosts and their degree
of immune suppression [9], it is possible that there may be an intrinsic path-obiologic varietal difference, particularly in the propensity to invade the
brain parenchyma.
Skin
Cryptococcus neoformans has been reported to produce almost any
type of human skin lesion [144–147]. Skin manifestations include acneiform
or endocarditis) but these conditions have been reported. Bone and jointinvolvement is an occasional extraneural site. Before the HIV epidemic, it
had been reported that up to 5% of disseminated cases of cryptococcosis
showed bone and joint involvement [171]. Osteoarticular cryptococcosis can
occur in both immunocompromised and nonimmunocompromised hosts
and the only clinical link with infection is the suggestion that sarcoidosis
is a common underlying disease with this site of infection. In AIDS patients,
it might be found in a random bone marrow biopsy without any radio-
graphic evidence of disease. Cryptococcal peritonitis is an occasional clinicaloccurrence and generally presents in two groups of patients: patients on
chronic ambulatory peritoneal dialysis; and patients with severe underlying
850 J.R. Perfect, A. Casadevall / Infect Dis Clin N Am 16 (2002) 837–874
illnesses, such as cirrhosis [172]. The following body sites are considered rare
manifestations of cryptococcosis because less than a dozen cases per site
have been presented in the literature: genitourinary tract (pyelonephritis andgenital lesions); muscle (myositis); heart (native and prosthetic endocarditis,
mycotic aortitis or aneurysm, myocarditis, pericarditis, and vascular foreign
body); thyroid (thyroiditis and mass); adrenal gland (adrenal insufficiency
and excess); head and neck (gingivitis, sinusitis, salivary gland involvement,
larynx, and neck mass); gastrointestinal (esophagitis, biliary tract, enteritis,
and hepatitis); breast (mastitis and mass); and lymph node (lymphadenop-
athy) [46].
Laboratory diagnosis
Direct microscopic examination
Clinical specimens from the CSF to aspirates of cutaneous lesions can be
examined by India ink examination for the 5 to 10 lm in diameter yeast cells
with capsules. Approximately 80% of AIDS patients and 50% of non-AIDS
patients with cryptococcal meningitis reveal the yeast with this simple and
immediate test. For a routine positive India ink test from CSF, it is esti-mated that the specimen needs to contain greater than 103 CFU/mL of CSF
(personal observations). Although calcofluor white is not specific for C. neo-
formans, this solution can be used on tissue aspirates with a fluorescent
microscope to find these yeasts quickly when they are in a reduced number
within a specimen. In tissue sections, the encapsulated yeasts may be sur-
rounded by empty spaces because the capsule does not routinely stain with
many of the standard histologic stains, such as hematoxylin and eosin. The
appearance of the polysaccharide capsule can be identified with specificstains, such as mucicarmine and alcian blue. The Gomori methenamine sil-
ver stain for fungi identifies C. neoformans as a yeast but it is not specific for
this yeast and the Fontana-Masson stain, which identifies melanin, selec-
tively stains these yeasts in tissue. Most clinical isolates in both tissue and
fluids have evidence of a capsule, although occasionally there are reported
hypocapsular strains in which the capsule is difficult to observe histologi-
cally [173].
Cultures
Cryptococcus neoformans can be isolated on most routine mycologic or
bacteriologic media. Standard blood culture methods, including radiometric
methods, now routinely detect cryptococcemia. In fact, one study using a
lysis-centrifugation procedure showed over 70% sensitivity of detecting
cryptococcemia in AIDS patients with cryptococcosis [174]. Most C. neofor-mans isolates can be detected in culture between 3 and 7 days after the speci-
men has been placed into or on media.
851J.R. Perfect, A. Casadevall / Infect Dis Clin N Am 16 (2002) 837–874
All Cryptococcus species are nonfermentative and produce extracellular
starch and urease. C. neoformans is unique among Cryptococcal species in
its ability routinely to grow at 37�C. It also produces a phenoloxidase. Thesefeatures have allowed for the rapid identification of C. neoformans by the
use of a rapid urease test or the use of Staib’s birdseed agar, DOPA, or caf-
feic acid media to detect the brown or black colonies as the phenoloxidase
breaks the diphenolic compounds in the media down into melanin [175–
177]. Isolates can also be identified by several commercial systems based
on biochemical reactions [178,179] or the use of DNA-based methods
[180,181].
The two varieties of C. neoformans can be differentiated by a color reac-tion when grown on concanavine-glycine thymol blue agar [182] and a sero-
type commercial kit using monoclonal antibodies is available [183,184].
Finally, specific strains can be fingerprinted or biotyped using DNA-based
or enzymatic methods, respectively [185–188], and these DNA methods can
identify specific varietal genotypes.
Serology
The serologic test for the detection of cryptococcal polysaccharide and
the diagnosis of invasive cryptococcosis is both specific and sensitive
(�90%). The most commonly used method of antigen detection is the latex
agglutination test, but the enzyme immunoassay is occasionally used in
some medical centers. A positive test at titers of greater than or equal to
1:4 in a biologic fluid strongly suggests infection. In fact, every positive test,
no matter what the titer, should be considered carefully within the clinical
context of the patient. Many kits can now detect as little as 10 ng/mL of poly-saccharide in specimens. With proper treatment of specimens, such as elim-
ination of rheumatoid factor by boiling and pronase or 2-mercaptoethanol
treatment, there are few false-positives. One rare false-positive occurs when
there is a cross-reactive antigen, such as the polysaccharide of Trichosporon
beigelii, in the specimen or another microorganism [189–191]. False-negative
results, however, are also rare. These false-negative tests may be caused by
low titers, early infection, presence of immune complexes, prozone effect of
high titers, or poorly encapsulated strains with low production of polysac-charide. The antigen detection system has its most clinical experience with
CSF and serum specimens, but it has been used to detect polysaccharide
antigen in urine or bronchoalveolar lavage fluid [192].
In diagnosis, cryptococcal antigen tests should be part of the standard
analysis on CSF specimens in immunocompromised hosts with meningitis.
They may also be used to screen serum in febrile, very high-risk patients,
such as those with HIV infection in areas where there is a known high rate
of cryptococcal infection [193]. The detection of cryptococcal polysacchar-ide is the leading example of a useful test for serologic diagnosis of an inva-
sive fungal infection and it may even be positive before detection of the
852 J.R. Perfect, A. Casadevall / Infect Dis Clin N Am 16 (2002) 837–874
organism by culture. Despite its excellence as a diagnostic test, it is more dif-
ficult to use as a treatment barometer except as a general prognostic feature.
High titers (�1:1024) generally reflect high burden of yeasts, a poor hostimmune response, and a greater likelihood of therapeutic failure. Crypto-
coccal antigen titers have been less useful to make specific clinical treatment
decisions. For instance, the use of serial antigen titers to develop a treatment
algorithm remains imprecise and not validated by clinical trials.
Antibodies to C. neoformans may not be detected or are at low levels dur-
ing active infection because many patients are severely immunosuppressed,
but they may rise during clinical recovery. They are not useful, however, for
diagnosis or treatment decisions.
Radiology
Chest radiograph
The chest radiograph of pulmonary cryptococcosis in the immunocompe-
tent host can show a variety of features including nodules, infiltrates, hilar
lymphadenopathy, or pleural effusions [136,194–198]. In AIDS patients the
radiograph may reveal either diffuse or focal interstitial infiltrates with orwithout lymphadenopathy [124,199,200]. This radiograph may be confused
with P. carinii infection or represent co-infection with it or another
pathogen.
CT and MRI
CT and MRI are frequently used in the diagnosis and management of
cryptococcal meningitis. CT scan findings in non-AIDS patients with men-
ingitis can reveal hydrocephalus; gyral enhancement; or multiple nodules,which may be enhancing or nonenhancing [201]. Cryptococcomas can be
either single or multiple and occur in up to 25% of patients [202,203].
Approximately one half of CT scans, however, are normal in cryptococcal
meningitis. In patients with HIV infection, the scans are similar to non-
AIDS patients except approximately one third of patients have cortical atro-
phy from their underlying HIV infection [204].
The MRIs are more sensitive than CT scan for detecting abnormalities.
MRI findings include numerous clustered foci that are hyperintense onT2-weighted images and nonenhancing on postcontrast T1-weighted images
within basal ganglia and mid-brain, which represent Virchow-Robin spaces.
There may also be the multiple miliary enhancing parenchymal and lepto-
meningeal nodules with gadopentetate dimeglumine [205].
It should be emphasized that there is no pathognomonic radiographic
picture for cryptococcal meningitis. It may present simply as an idiopathic
hydrocephalus identified by CT or MRI [206]. In AIDS patients, however,
a scan may identify a parenchymal lesion, which represents a second CNSdisease process, such as toxoplasmosis or lymphoma. Finally, the use of
CT or MRI in follow-up of cryptococcal meningitis may show worsening
853J.R. Perfect, A. Casadevall / Infect Dis Clin N Am 16 (2002) 837–874
(enlargement or new lesions) or persistence of the cryptococcomas [207]
while the patient continues to improve clinically or is stable. This finding
may simply represent enhancement by inflammatory cells within granulo-mas as microscopic infectious foci are being eliminated; it is not necessarily
a criteria for treatment failure.
Management
General
The management of cryptococcosis has been extremely well-studied and
although all aspects of treatment are not consistently successful or uni-
formly agreed on, there is substantial evidence-based data to make recom-
mendations. A group of experts have attempted to collate the therapeutic
evidence into a series of guidelines for therapy of cryptococcosis [208]; these
guidelines are excellent starting points for therapeutic decisions, but this dis-
cussion adds the authors’ own insights and opinions. The breadth of deci-
sions to be made in cases of cryptococcosis can be quite diverse. Forinstance, it is clear that some individuals with asymptomatic isolation of
C. neoformans from respiratory secretions have been untreated without clin-
ical compromise. A recent review of this issue suggested that approximately
20% of patients with positive pulmonary cultures received no treatment
[121]. Cryptococcal meningitis, however, is uniformly fatal without antifun-
gal treatment. Before availability of antifungal agents, there were cases that
survived for years with this chronic CNS condition before succumbing. The
rapidity of progression to death for the natural history of this infection withsevere immune suppression, however, such as HIV infection, is illustrated
in the comprehensive report on meningitis in HIV-infected patients in
Zimbabwe [72]. Of 406 patients, 45% had cryptococcal meningitis and
almost 40% of those with this infection died during the initial hospitalization
without treatment. Therapeutic decisions do have a wide range of options
and consequences in the management of cryptococcosis.
In vitro susceptibility testing
Methods for testing C. neoformans isolates in vitro for minimum
inhibitory concentrations (MICs) have been standardized and modified for
media, inoculum, and end point determinations [209,210]. Most initial
isolates of C. neoformans have lowMICs to amphotericin B, flucytosine, and
azoles [211,212]. Initial resistance to flucytosine is low [213] but rising MICs
occur during therapy with this agent and correlate with clinical relapse
[137,214,215]. There has been identified, however, only an occasional
C. neoformans isolate resistant to in vitro amphotericin B [216], andmost relapse isolates remain fully susceptible to this polyene. During the
HIV epidemic and before HAART the widespread use of azoles in severely
854 J.R. Perfect, A. Casadevall / Infect Dis Clin N Am 16 (2002) 837–874
immunocompromised host showed an increase in lumenal infections with
azole-resistant Candida albicans. Although there have been some apparently
azole-resistant C. neoformans strains isolated from patients, azole resistanceprobably runs less than 5% of initial isolates [217]. In fact, when secondary
isolates are examined, it seems through molecular typing methods that these
are relapse isolates rather than reinfection [218] and most have a similar (sus-
ceptible) MIC as the primary isolates [219]. There are cases of increasing flu-
conazole MICs during treatment [220], however, and present animal data
suggest a correlation between in vitro susceptibility testing and in vivo out-
come [221]. It is not certain what the precise breakpoint is for fluconazole sus-
ceptibility in cryptococcosis treatment. MICs, however, of greater than orequal to 16 lg/mL from both pharmacodynamic studies in vitro [222] and
some experience with isolates from relapsed patients with prior azole expo-
sure [223] should make the clinician consider very high dosing of drug or the
possibility of using another agent. It is possible that more azole-resistant C.
neoformans with its heteroresistant phenotypes [224] may be observed in the
future. At this time with the low incidence of primary azole-resistant isolates
it is probably not necessary to check MICs before treatment if patients had
not received prior azole therapy. All initial isolates should be saved and com-pared with relapse isolates, however, to determine if drug resistance is a pos-
sible factor for treatment failure. A general overview of in vitro and in vivo
reports suggests that there is a positive interaction against C. neoformans for
the following drug combinations: amphotericin B and flucytosine; amphoter-
icin B and azole; amphotericin B and rifampin; flucytosine and azole; and tri-
ple combination of amphotericin B, flucytosine, and azole [225–227].
Treatment strategies
It should be emphasized that success rates from any study depend pri-
marily on study definitions and the patient’s prognostic factors. These fac-
tors always need to be taken into account when reviewing the published
data and treating the individual patient.
Amphotericin B therapy converted cryptococcal meningitis from a uni-
formly fatal infection to one that is curable. Amphotericin B successes
before the AIDS epidemic were reported at 60% to 70% [146,228]. A 10-week course of amphotericin B at 0.4 mg/kg/d for 10 weeks recorded a suc-
cess rate of 68% in non-AIDS patients [228]. In AIDS patients at 0.5 mg/kg/
d of amphotericin B, the success rate dropped to 40% [229]. Recent studies
have suggested that a higher daily dose of amphotericin B may be more
effective [230,231] and this strategy has been more commonly adopted. Fur-
thermore, amphotericin B can now be given in higher doses with reduced
toxicity in lipid preparations if toxicities develop with standard amphoteri-
cin B. Ambisome at 4 mg/kg/d has the most clinical experience [3] andincludes a comparative trial in which it performed similar to amphotericin
B [232]. It is probably the best alternative polyene preparation for treatment
855J.R. Perfect, A. Casadevall / Infect Dis Clin N Am 16 (2002) 837–874
after standard amphotericin B deoxycholate, although ABLC has shown
some treatment success in cryptococcal meningitis [233].
Flucytosine was initially used alone in the treatment of cryptococcal men-ingitis [214,234], but development of resistance on therapy reduced enthusi-
asm for monotherapy. Several studies used the combination of amphotericin
B (0.3 mg/kg/d) and flucytosine (150 mg/kg/d) to reduce the course of treat-
ment to 6 weeks and the toxicity of amphotericin B [228,235,236]. In some
cases, with good prognostic signs there was success with 4 weeks of treat-
ment [236]. Furthermore, in a small number of AIDS patients, the combina-
tion therapy was found to be superior to fluconazole [237] and itraconazole
alone [238]. The combination regimen reproducibly sterilizes CSF at 2 weeksin most patients including those with AIDS and there are several studies that
support less relapses with an amphotericin B plus flucytosine treatment reg-
imen [239,240]. This led to a definitive study using higher doses of ampho-
tericin B (0.7 mg/kg/d) and lower doses of flucytosine (100 mg/kg/d) for a
2-week induction period of treatment and then switching to fluconazole
(400 mg/d) alone for 8 weeks and completion of initial therapy [241]. This
strategy has become a primary modality in AIDS patients and frequently
chosen now for non-AIDS patients with cryptococcal meningitis [121] andin the authors’ opinion represents the initial regimen of choice for treatment
of cryptococcal meningitis.
Fluconazole has been studied in cryptococcal meningitis because of its
excellent pharmacokinetics within the CNS. In a comparative trial with
amphotericin B in AIDS patients, the success rate was similar to amphoter-
icin B alone but sterilization of the CSF took longer with the azole therapy
[229]. Fluconazole may have its most difficult time in controlling infection
when the burden of yeasts is high and infection is well-established[242,243]. In initial therapy of cryptococcal meningitis it may be less useful.
Fluconazole has also been used as salvage therapy with higher doses [244].
Furthermore, it has been used successfully with flucytosine in treatment of
an animal model [245] and humans [246]. It has also been used in successful
suppressive therapy for AIDS patients [247,248] and may be useful for pro-
longed initial therapy in non-AIDS patients. Although optimal dosing for
initial stages of meningitis treatment is not precise, doses of 400 to 800
mg/d are likely to give the best results.Itraconazole, despite its poor penetration into CSF, has been used in the
successful management of cryptococcal meningitis [249–251]. Direct com-
parison for initial treatment of cryptococcal meningitis with fluconazole has
not been made. For suppressive therapy in AIDS patients, however, fluco-
nazole has been shown to have a distinct advantage over itraconazole [239].
Miconazole has rarely been used in cryptococcal infection and with
mixed results [252–254]. Ketoconazole has not been successful in treatment
of cryptococcal meningitis [255], but it is likely to be effective in non-CNSinfections [256]. Voriconazole and posaconazole have excellent in vitro
activity against C. neoformans and animal studies support their potential
856 J.R. Perfect, A. Casadevall / Infect Dis Clin N Am 16 (2002) 837–874
in this infection [257,258]. Present trials for these drugs in cryptococcosis
have generally used them as therapy for patients who failed standard regi-
mens. Successes in this unique population of patients run from 30% to70% [259,260]. At present, the new b-glucan synthase inhibitors, such as cas-
pofungin, micafungin, and anidulafungin, do not have clinical activity for
treatment of C. neoformans. There are a series of concepts in treatment of
cryptococcosis that require further explanation.
Site of infection
In pulmonary cryptococcosis, there are few comparative studies to deter-
mine which drugs are the best when treatment is indicated. Azoles, however,
such as fluconazole, at 200 to 400 mg/d for 3 to 6 months, are safe and easy
to administer and generally form the primary treatment strategy [261,262],
unless the patient is severely ill, in which case amphotericin B should beused. In the authors’ opinion, it is clear that both symptomatic and asymp-
tomatic immunosuppressed patients with C. neoformans isolated from lung
must be treated because dissemination from this site is a distinct possibility
[137]. In fact, the authors recommend that most patients irrespective of
immune status should probably be treated when C. neoformans is isolated
from nonsterile respiratory secretions with or without symptoms. Most CNS
cryptococcomas can be treated and although length of therapy is not
defined, the authors treat patients sometimes with 2 years of fluconazole.As previously mentioned, serial scans may not be helpful and complete res-
olution radiographically may take years. Surgery has been recommended for
large CNS lesions (>3 cm) [263], but in the authors’ opinion its use needs to
be individualized, with it rarely needed with present medical management.
Immune status of host
Every attempt to improve immunity of the host during treatment needs to
be made. This may mean reducing corticosteroid treatment and it is recom-
mended that a dose of prednisone less than or equal to 20 mg/d be achieved
during therapy. More studies on cytokines as adjunct therapy are needed(see later). The published results of these studies are awaited to attempt to
put cytokine treatment in proper perspective. Unfortunately, in AIDS
patients who generally develop C. neoformans with less than 100 CD4
cells/lL, immune augmentation had been difficult until the age of HAART
and continuous azole suppressive therapy had been required to prevent a
high rate of relapse (50% to 60%) if antifungals were stopped, but this strat-
egy may be changing (see later). In HIV-infected patients with cryptococcal
meningitis, it may be reasonable first to treat the fungal infection aggres-sively and attempt to sterilize CSF with antifungal drugs before starting
HAART to avoid a rapid inflammatory process return and development
857J.R. Perfect, A. Casadevall / Infect Dis Clin N Am 16 (2002) 837–874
of new symptoms. HAART can probably be started at 2 to 3 months after
initiation of specific anticryptococcal therapy.
Suppressive therapy
Relapse rates in AIDS patients with cryptococcal meningitis before
HAART approached 50% to 60% when antifungal therapy was stopped.
Studies showed that daily fluconazole can reduce relapse rates to less than5% [248,264]. Fluconazole has been shown to be more effective than weekly
intravenous amphotericin B dosing [248] and itraconazole [239] and it has
become standard clinical practice to prescribe indefinite fluconazole sup-
pressive therapy in AIDS patients. Recent data in several small studies,
however, suggest that with HAART and its immune reconstitution (rising
CD4 counts and lowering HIV loads), antifungal suppressive therapy can
be stopped at 1 year with little chance for relapse in the next year [265–
267]. A large study of stopping the suppressive azole regimen is beginningin Europe and it is hoped this study will bring more certainty to this strat-
egy. There are no guidelines for suppressive therapy in non-AIDS patients
with cryptococcal meningitis who can relapse in the first year at a 10% to
15% rate with standard regimens. With the use of safe and oral azoles, a
common practice has been to extend therapy with fluconazole at 200 mg/d
for 6 months to a year to cover this highest relapse period.
Role of intracranial pressure
Patients with high burden of yeasts and antigen can develop increased
intracranial pressure both before treatment, during early treatment, and
with the chronic phase of treatment and monitoring. With high intracranialpressures during early management of infection, these patients develop
symptoms, such as systemic hypertension, decreased sensorium, cranial neu-
ropathies, visual loss, or increased headaches. It has been postulated that the
elevated subarachnoid pressures are caused by reduced CSF resorption
because of increased outflow resistance possibly from yeasts and polysac-
charide plugging the arachnoid villi and the antigen contributing to brain
edema [268]. Clinical response to therapies is better in those patients with
pretreatment CSF opening pressures less than or equal to 250 mm H2Oor those with stable or decreased (>10 mm H2O) lumbar CSF pressures
at 2 weeks [269]. Control of increased intracranial pressures with techniques
of external drainage, such as repeated lumbar punctures or lumbar drains,
may be necessary during the early treatment phase. Unfortunately, the gen-
eral preciseness of methods and goals remains elusive and generally is based
on individual patient responses. In contrast to early treatment pressure
problems, ventricular shunts are successful for long-term pressure control
in those with classic obstructive hydrocephalus, which may occur monthsafter start of treatment and must be watched for during follow-up visits.
A shunt can be placed successfully while cryptococcal meningitis is being
858 J.R. Perfect, A. Casadevall / Infect Dis Clin N Am 16 (2002) 837–874
effectively treated. In cases of cryptococcal meningitis in which the shunt has
already been in place for treatment of hydrocephalus before anticryptococ-
cal treatment, however, it needs to be removed for cure [206]. The patientcan then be started on adequate treatment and when CSF is sterile another
shunt can be placed if clinically indicated.
Antigen titers
Cerebrospinal fluid polysaccharide antigen titers generally reflect the bur-
den of organisms at the initial diagnosis and high antigen titers predict apoor prognosis. The use of specific antigen titers to predict individual out-
come and direct therapy, however, is generally not helpful [270]. Polysac-
charide antigen may persist for long periods of time despite adequate
therapy. Changing or fixed titers are not reliable indicators of treatment out-
come in individual patients. During the AIDS epidemic, isolated cases of
serum cryptococcal polysaccharidemia in HIV-infected patients with nega-
tive cultures have occurred [271]. Management of these cases is difficult but
empiric therapy may be wise in these high-risk patients because many even-tually develop cryptococcosis.
Cytokines and monoclonal antibodies
Disseminated cryptococcosis generally occurs during an episode of
immunosuppression. There are in vitro studies and animal models that indi-cate that biologic response modifiers or antibodies can have a positive effect
on outcome of cryptococcosis (see host factors). Granulocyte-macrophage
IL-12, and interferon-c can positively interact with host cells to inhibit or
kill C. neoformans. Granulocyte-macrophage colony–stimulating factor has
been used in an open trial [272] and results of an adjunct trial of interferon-
c, which was supported by encouraging results in animals, are awaited [273].
It is not yet clear whether these cytokines help or exactly how to optimizetheir use. Specific monoclonal antibodies and serum therapy have a positive
effect on the prevention and treatment of experimental cryptococcosis [274–
276]. A monoclonal antibody is now being studied in early phase I trials in
HIV-infected patients with stable cryptococcal disease but persistent poly-
saccharide antigenemia. There remain little definitive human clinical trials,
however, into immune modulation therapy. It is clear that future studies
with these biologic enhancers are needed as further strategies are developed
for management in an infection, which can still have 10% to 25% acute mor-tality rates during early treatment in medically advanced countries.
Prognosis
The most important prognostic factor for the success and treatment
of cryptococcal meningitis is the ability to control the patient’s underlying
859J.R. Perfect, A. Casadevall / Infect Dis Clin N Am 16 (2002) 837–874
disease. It has been shown that cancer victims have a shorter survival than
AIDS patients because of their underlying disease state [277]. Another major
risk group is solid organ transplant recipients and recent data and prognosisin this group are conflicting. In one study the prognosis of cryptococcosis in
solid organ transplant was similar to patients without an underlying disease
[121] but in a second study there was a death rate of 42% [43]. More studies
are needed in this area. Two other prognostic findings are burden of organ-
isms at presentation and the level of patient’s sensorium. A poor prognosis is
found with a heavily positive India ink examination; high polysaccharide
antigen titers (�1:1024); and a poor inflammatory response in the CSF
(<20 cells/ll). Patients on presentation who have a lucid sensorium have abetter prognosis than those stuporous or in coma [236].
Specific factors have predicted outcome and generally relate to the three
factors previously described. On amphotericin B therapy, Diamond and
Bennett [278] found more than three decades ago that patients who died
during therapy were more likely to have: (1) an initial positive India ink
CSF leukocytes, (5) cryptococci isolated from extraneural site, (6) absence of
anticryptococcal antibody, (7) initial CSF or serum antigen titer of greaterthan 32, and (8) corticosteroid therapy or lymphoreticular malignancy.
Patients who relapsed after treatment were characterized by: (1) abnormal
CSF glucose for greater than or equal to 4 weeks of therapy, (2) low initial
CSF leukocytes, (3) cryptococci isolated from extraneural sites, (4) absence
of anticryptococcal antibody, (5) posttreatment CSF or serum cryptococcal
antigen titer greater than or equal to 8, (6) no significant decrease in CSF
and serum antigen titer during therapy, and (7) daily dose of corticosteroid
therapy equivalent to 20 mg of prednisone or more after therapy. Theseinsights may still help to predict failures today. Recent studies with ampho-
tericin B and flucytosine treatment in non-AIDS patients indicated a better
prognosis if there was a normal mental status, headache on presentation,
and a CSF leukocyte count greater than 20 WBCs/lL [236]. In AIDS
patients during treatment with amphotericin B or fluconazole, important
positive pretreatment predictors for death during treatment were abnormal
mental status, a CSF antigen titer greater than 1:1024, and CSF leukocyte
count less than 20 WBC/lL [241]. Identification of these high-risk patientsfor failure and relapse should allow the clinician to design specific antifungal
regimens for these subsets of patients.
Prevention
There are at least four methods for approaching high-risk patients to pre-
vent cryptococcosis. First, antifungal prophylaxis for AIDS patients has
been effective in reducing the incidence of cryptococcosis with the use of flu-conazole [67,279]. Both HAART and concerns about widespread exposure
to azoles worked together, however, to reduce enthusiasm for its general
860 J.R. Perfect, A. Casadevall / Infect Dis Clin N Am 16 (2002) 837–874
use. Second, active immunization with a vaccine for high-risk patients is
possible. A cryptococcal GXM-tetanus toxoid conjugate vaccine has been
developed which elicits protective antibodies in mice [280] and an initialparadigm has been completed. It or other vaccine candidates, however,
await clinical trials in humans. Third, the ability to produce protective
humanized or murine monoclonal antibodies could be given to high-risk
patients as passive prophylactic therapy, but protection requires repeated
continuous administration over the risk period. Finally, high-risk patients
can attempt to avoid high-risk environments, such as bird droppings, which
contain yeasts to be aerosolized and inhaled.
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